Microparticles from streptococcus pneumoniae as vaccine antigens

ABSTRACT

An isolated Streptococcus pneumoniae membrane vesicle microparticle (MP), wherein said MP comprises: the protein Ply at the level of ≥0.070 μg/μg total protein in the MP; and/or the protein LytA at the level of ≥0.070 μg/μg total protein in the MP; and/or the protein PspC at the level of ≥0.130 μg/μg total protein in the MP; and/or the protein RrgB at the level of ≥0.020 μg/μg total protein in the MP. Compositions comprising such MPs. Uses thereof in particular in immunization, as well as methods of manufacture thereof.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.16/474,940, filed on Jun. 28, 2019, which is a National Stageapplication under 35 U.S.C. § 371 of International Application No.PCT/SE2017/051323, having an International Filing Date of Dec. 21, 2017,which claims the benefit of SE Application No. 1651746-8 filed Dec. 28,2016. The contents of the aforementioned applications are herebyincorporated by reference in their entirety.

TECHNICAL FIELD

The present invention relates to the field of vaccines againstStreptococcus pneumoniae.

BACKGROUND TO THE INVENTION

Streptococcus pneumoniae is a Gram-positive bacterium that is a majorcontributor to morbidity and mortality worldwide causing about 2 milliondeaths annually, i e the same range as for tuberculosis. Pneumococci arethe major cause of common infections such as sinusitis, otitis, andcommunity-acquired pneumonia (CAP), but also a common cause of severeinvasive diseases (IPD) such as sepsis and meningitis. Young childrenand the elderly are the most susceptible age groups to acquirepneumococcal infections, but other factors such as underlying diseases,splenectomy, immunosuppression (including HIV), diabetes and a priorinfluenza-virus infection also sensitize for a pneumococcal infection.Pneumococcal infections such as CAP also predispose for cardiovasculardiseases such as cardiac infarction, and meningitis patients may getneurological sequelae such as hearing loss and cognitive impairments.

Pneumococci have the human upper airways as their normal ecologicalniche, and up to 60% of preschool children may be colonized at any giventime without symptoms. It is believed that most transmission events tothe elderly come from young colonized children. S. pneumoniae is ahighly genetically diverse species due to an efficient DNA exchangesystem resulting in a multitude of clonal lineages that may differ fromone another in the presence and absence of genes and gene clusters, aswell as in variations in individual genes.

Variations in the capsular locus, the major virulence factor and vaccinetarget of pneumococci, have resulted in at least 97 known capsularserotypes with different potential of causing disease.

Pneumococcal conjugate vaccines (PCVs), targeting a limited number (7,10 or 13 in PCV7, PCV10 and PCV13 respectively) of the known 97 capsularserotypes, have been introduced in the childhood vaccination program inmany countries. Vaccine introduction has led to a decrease in IPD causedby vaccine-types in vaccinated children, however, non-vaccine typepneumococci have rapidly expanded and replaced vaccine-types inchild-hood carriage and in IPD, but also in non-vaccinated populationsuch as the elderly. Furthermore, protection against vaccine typesdiffer, and for serotype 3, a serotype included in PCV13 and with a highmortality rate, there is more or less no evidence for protection againstIPD so far. Data from Sweden, where both PCV10 and PCV 13 are used, showthat serotype 3 was a dominating serotype among IPD cases during 2016,also in counties using only PCV13. Hence, novel vaccine approaches areneeded.

In Sweden the total number of patients with IPD has remained almost thesame after vaccine introduction. Moreover, in a recent study we foundthat pneumococcal carriage rates remained around 30% in the youngestchildren 8 years after vaccine introduction in Stockholm, andnon-vaccine types were found in over 90% of the isolates. Antibioticresistance is emerging among clinical pneumococcal isolates andaccording to reports from the European Centre for Disease prevention andControl (ECDC), and the European Antimicrobial Resistance SurveillanceNetwork (EARS-Net), reduced susceptibility to penicillin has increasedduring the last years in Sweden and now exceeds 5% among IPD cases. Inaddition, the need for multiple antigens covering various capsularserotypes lead to high production costs for the known vaccines.

Opsonophagocytosis is the primary mechanism for clearance of pneumococcifrom the host, and the measurement of opsonophagocytic antibodiesappears to correlate with vaccine-induced protection. However, it hasproven difficult to design vaccines that elicit an efficient response inform of opsonophagocytic antibodies.

Taken together, the above issues stress the need for novel vaccineapproaches. Thus, an object of the present invention is the provision ofimproved pneumococcal antigens able to confer serotype-independentimmunological responses and/or improved response in terms ofopsonophagocitic antibodies and protection in experimental models.

Background for Pneumococcal Microparticles

Spherical membranous blebs, so called outer membrane vesicles (OMVs),are produced by Gram-negative bacteria. OMVs range in size from 10 to300 nm and they have been shown to be formed by budding from thebacterial outer membrane, and to have many functions, such asinfluencing virulence by different mechanisms. In Gram-negative bacteriaOMVs have been suggested to act as vehicle to deliver virulence factorsto host cells. Only recently, membrane vesicles (MV_(L)) were discoveredalso in Gram-positive bacteria and their formation and function ispoorly understood compared to OMVs. Gram-positive bacteria do notcontrain an outer membrane and the the cell wall differs fromGram-negative bacteria.

For Streptococcus pneumoniae MV_(L) (membrane vesicle) were isolatedfrom pneumococci belonging to different serotypes (2, 6B, 8 and 23F)grown in liquid culture medium (1). MV_(L) from serotype 8 were shown tobe protective against pneumococcal challenge with the same serotype inmice. Importantly, no cross protection was shown to pneumococci of otherserotypes.

Definitions

The pneumococcal capsule. The pneumococcal capsule, consisting ofpolysaccharide, exists in at least 97 different variants, so calledcapsular serotypes. The pneumococcal capsule is a major virulencedeterminant of pneumococci, affecting phagocytosis of the bacteria byhost cells, and influence bacterial interactions with the innate immuneresponse.

Pneumolysin (termed Ply herein) is a 53 kDa cholesterol dependentcytolysin released by Streptococcus pneumoniae upon lysis. It is one ofthe major virulence factors of this bacterium. It forms pores in alleukaryotic cells that have cholesterol in their membranes. The formationof pores by Ply frequently results in host cell death as membraneintegrity is destroyed. Ply plays a central role in protecting thepneumococcus from complement attack and aiding its spread to othertissues/organs. Ply is able to activate the classical complementpathway, even in the absence of Ply specific antibody (2). A referencesequence from strain TIGR4 is presented in SEQ ID NO: 1.

LytA is the major autolysin of Streptococcus pneumoniae. Lysis is causedby cleaving the lactyl-amide bond between the stem peptides and theglycan strands of peptidoglycan, resulting in hydrolysis of the cellwall. The contribution of LytA to pneumococcal virulence is stillunclear. It is possible that LytA-mediated lysis releases othervirulence factors such as pneumolysin. LytA could also be released tolyse neighboring non-competent pneumococcal cells in a fratricidalmanner. This would potentially facilitate genetic exchange betweennaturally competent pneumococcal populations that easily take up andincorporate DNA by homologous recombination. A third possibility is thatLytA mediates lysis to release proteins involved in immune evasion orcell wall components that may interfere with the host immune response(3). A reference sequence from strain TIGR4 is presented in SEQ ID NO:2.

PspC/CbpA (choline binding protein A) is a protein that binds thephosphocholine present in the teichoic acid and the lipoteichoic acid ofthe cell membrane and the cell wall. It is a major pneumococcal adhesin.It promotes pneumococcal adherence via a human-specific interaction withthe ectodomain of the polymeric Ig receptor. It also prevents activationof C3b and complement-mediated opsonophagocytosis of pneumococci (4).The pspC locus is highly polymorphic and 11 major groups of this proteinhave been identified. Single PspC proteins are identified by sequentialnumbers separated from the group number by a dot (5) as follows:PspC1.1, PspC2.1, PspC2.2, PspC3.1, PspC3.4, PspC4.2/PspC10.1, PspC5.1,PspC6.1/PspC9.1, PspC7.1, PspC8.1, PspC11.1 and PspC11.4 (SEQ ID NOs:3-14, respectively).

RrgB is the major subunit and stalk protein of the pneumococcal pilus.S. pneumoniae pilus 1 is encoded by a genetic islet (PI-1) present in 30to 50% of the pneumococcal strains and is implicated in adhesion toepithelial cells, lung infection, and virulence. Pilus 1 is composed ofthe backbone subunit RrgB, the minor pilin subunits RrgA, and RrgC (6).A reference sequence from strain TIGR4 is presented in SEQ ID NO: 15.

RrgA is a minor pilin subunit of the pneumococcal pilus. RrgA is the tipprotein of the pilus and has been shown to mediate adhesion toepithelial cells (6, 7). Recently, RrgA was shown to promote meningitisdevelopment in mice (8). A reference sequence is presented in SEQ ID NO:16.

PhtD is a polyhistidine triad protein of S. pneumoniae. Polyhistidinetriad (Pht) proteins are streptococcal surface proteins that containmultiple copies of a characteristic HxxHxH sequence, designed ashistidine triads, which were predicted to bind divalent metal cations.Despite the increasing amount of biochemical, structural andphysiological data, the functional role of PhtD and the other Phtproteins remains unclear. PhtD could be involved in zinc uptake.Alternatively, Pht proteins could play a role in protecting thepneumococcus from toxic effects of high Zn²⁺ concentrations byscavenging, storing or trapping Zn²⁺ ions (9). A reference sequence fromstrain TIGR4 is presented in SEQ ID NO: 17.

IgA refers to Immunoglobulin A1 protease of S. pneumoniae (10). SaidIgA1-protease is a highly specific enzyme that cleaves amino acidsequences of certain proteins. The natural substrate of the IgA proteaseis immunoglobulin A1, hence its name. The enzyme is in fact capable ofcleavage of proteins with the amino acid sequenceN-X-Z-Pro-Pro/-Y-Pro-C, where the X in the sequence is preferably aProline or Serine; the Y=Threonine, Serine or Alanine; and Z preferablyis Arginine or Threonine. Thus, the IgA1 protease acts by cleaving theproline-rich hinge region of the heavy chain of IgA1. Release of theIgA1 protease by the pathogen allows adherence to mucous membranes bydegrading host immunoglobulin A. A reference sequence from strain TIGR4is presented in SEQ ID NO: 18.

The term protective immunity in the present context refers toimmunization measures resulting in any degree of reduction in thelikelihood of developing the condition for which the protective immunityis relevant, including a minor, substantial or major reduction inlikelihood of developing the condition as well as total prevention.Preferably, the degree of likelihood reduction is at least a minorreduction.

The term sequence identity expressed in percentage is defined as thevalue determined by comparing two optimally aligned sequences over acomparison window, wherein a portion of the sequence in the comparisonwindow may comprise additions or deletions (i.e., gaps) as compared tothe reference sequence (which does not comprise additions or deletions)for optimal alignment of the two sequences. The percentage is calculatedby determining the number of positions at which the identical amino acidresidue occurs in both sequences to yield the number of matchedpositions, dividing the number of matched positions by the total numberof positions in the window of comparison and multiplying the result by100 to yield the percentage of sequence identity. Unless indicatedotherwise, the comparison window is the entire length of the sequencebeing referred to. In this context, optimal alignment is the alignmentproduced by the BLASTP algorithm as implemented online by the USNational Center for Biotechnology Information (see The NCBI Handbook[Internet], Chapter 16), with the following input parameters: Wordlength=3, Matrix=BLOSUM62, Gap cost=11, Gap extension cost=1.

BRIEF DESCRIPTION OF THE FIGURES

FIGS. 1A through 1D: Membrane-derived vesicles and microparticlesproduced by S. pneumoniae TIGR4 (T4). (FIG. 1A) Electron micrographs andatomic force micrographs (insets) of membrane vesicles andmicroparticles after isolation and density gradient purification fromliquid culture (MV_(L)) and microparticles from plate-grown (MP)bacteria. (FIG. 1B) Size distribution of MV_(L), MPs from TIGR4 and MPsfrom serotype 3. (FIG. 1C) Electron micrograph of a membrane buddingfrom S. pneumoniae T4R and (FIG. 1D) the same electron micrograph inlarger magnification. Released particle is indicated by arrow.

FIGS. 2A through 2D: Identification of proteins present in pneumococcalMV_(L) and MP. (FIG. 1A) Preparations of MV_(L) and MP were separated bySDS-PAGE and proteins visualized by coomassie staining. In comparison,proteins present in a whole cell lysate (WC) and supernatant (SN) of aliquid bacterial culture were visualized. (FIG. 1B) Venn diagramdisplaying the number of proteins identified by mass spectrometry(repeated twice) exclusively found in MV_(L) and MP preparations andcommon to both. (FIG. 1C) Mass spectrometry identification of proteinsfrom MV_(L) and MP. Numbers indicate percentages of proteins based ontheir subcellular localization, defined as cytosolic proteins,membrane-associated proteins (lipoproteins), proteins with onetransmembrane domain (N-terminally anchored), transmembrane proteins,cell wall associated proteins and secreted proteins. (FIG. 1D)Immunoblot detection of pneumococcal proteins and virulence factorspresent in MV_(L) and MP isolated from S. pneumoniae T4 WT, pneumolysin-(T4Δply) and LytA-deficient (T4ΔlytA) strains.

FIG. 3 : Pneumolysin associated with pneumococcal MV_(L) and MP isbiologically active. Quantification of in vitro hemolytic activity ofpurified MV_(L) and MP isolated from S. pneumoniae T4 and T4Δply.Erythrocytes from buffy coat blood were incubated for 60 minutes withdifferent concentrations (0.1, 0.8, 2, 6 and 10 μg protein) of MV_(L)and MP. As control treatments, erythrocytes were incubated with PBS (−)or 1% Triton X-100 (+). Data are represented as means+/−SEM of threeindependent experiments. *=P<0.05; **=P<0.01; ***=P<0.001.

FIGS. 4A through 4E: Microparticles (MP) are internalized by A549epithelial cells. (FIGS. 4A, 4B and 4C) Immunofluorescence pictures ofA549 cells treated with MP (10 μg/ml) for 24 hours. Cells were stainedfor F-actin, MP are detected with anti-pneumolysin and LytA antibodies.(FIG. 4A) Z-stack image (nr. stacks=21) of MP-treated cells.Internalized MP are indicated by arrows. (FIGS. 4B and 4C) Orthogonalviews of the same picture, taken at stack nr. 11, for the two indicatedMP, further showing their intracellular localization. (FIG. 4D)Immunoblot detection of anti-Ply, and anti-β-actin as loading control,in A549 cell lysates after incubation with increasing concentrations (1,2, 3 μg/ml) of MP. As control treatment, A549 cells were incubated withPBS (−). (FIG. 4E) Viability of A549 cells examined by flow cytometry offixable viability dye (FVD) positive cells after 24 hours of incubationwith MV_(L) or MP. As control treatment, A549 cells were incubated withPBS (−) or 0.02% NP40 in PBS. Data are represented as means+/−SEM ofthree independent experiments.

FIGS. 5A through 5C: MV_(L) and microparticles (MP) activate humanmonocyte-derived dendritic cells (DCs). (FIG. 5A) Percentages of FVD andAnnexin V-positive cells analyzed by flow cytometry. Cells wereincubated for 24 hours with different concentrations (10, 25, 50 μg/ml)of MV_(L), MP, PBS (−) or the unencapsulated mutant of TIGR4 (T4R). Dataare represented as means+/−SEM of three independent experiments.*=P<0.05. (FIG. 5B) Immunoblot detection of anti-Ply, and anti-S-actinas loading control, in cell lysates after incubation of DCs with MP (4μg/ml) in presence or absence of the inhibitors Cytochalasin D andWortmannin (C/W), to block active phagocytosis, or methyl-β-cyclodextrin(MβCD), to block lipid rafts and membrane fusion. Numbers represent therelative intensity of bands corresponding to Ply, adjusted to theloading control bands, in three independent experiments. (FIG. 5C) DCactivation measured by flow cytometry of MHCII and CD86-positive cellsafter 24 hours incubation with different concentrations (2.5, 5 μg/ml)of MV_(L), MP, PBS (−) or LPS (1 μg/ml). Data are represented asmeans+/−SEM of three independent experiments. *=P<0.05; ***=P<0.001.

FIGS. 6A through 6D: Cytokine release by human monocyte-deriveddendritic cells upon incubation with MV_(L) or MP. (FIG. 6A) IL-6, (FIG.6B) IL-8, (FIG. 6C) IL-10 and (FIG. 6D) TNF released by dendritic cellsafter 24 hours of incubation with different concentrations (10, 25, 50μg/ml) of MV_(L) or MP. Control treatments include PBS (−), theunencapsulated mutant of TIGR4 (T4R) and LPS (1 μg/ml). Not detectable(n.d.) amounts of cytokines are indicated. Data are represented asmeans+/−SEM of three independent experiments. *=P<0.05; **=P<0.01;***=P<0.001; ****=P<0.0001.

FIGS. 7A through 7E: Intranasal immunization of C57BL/6 mice with MV_(L)or MP of S. pneumoniae TIGR4 increase survival against intranasalinfection with serotype 1. (FIG. 7A) Percentage of mice survived afterchallenge. 20 mice per group. *=P<0.05; **=P<0.01; ****=P<0.0001. (FIG.7B) CFUs in lungs of mice after sacrifice. Each dot represents onemouse. **=P<0.01; ***=P<0.001. (C-E) CFUs in blood of mice. Each linerepresents one mouse.

FIGS. 8A through 8D: Intranasal immunization of mice with MV or MPsstimulate the production of pneumoccal-specific IgG. Detection ofMV_(L)-reactive (FIG. 8A), MP-reactive (FIG. 8B), T4-reactive (FIG. 8C)and T4R-reactive (FIG. 8D) mouse IgG in immunized mice sera using ELISAassay. Each dot represents one mouse serum. *=P<0.05; **=P<0.01;***=P<0.001; ****=P<0.0001.

FIGS. 9A and 9B: Staining of serotypes 1, 3 and 6B using antibodiesraised against MP from a serotype 4 strain. (FIG. 9A) Immunofluorescencepictures of pneumococcal strains belonging to serotypes 1, 3 and 6Bstained using sera, as primary antibodies, from mice immunized eitherwith MV_(L)+adjuvant or MP+adjuvant. The white spots on the bacterialsurface indicates the presence of specific antibodies againstpneumococci in the sera of the immunized mice. (FIG. 9B) Quantificationof the signal (white spots) detected on the bacteria; the signal ratiowas calculated by dividing the area of the signal detected on thebacteria (white spots) by the total area detected occupied by thebacteria. The area covered by the bacteria and the area covered by thesignal detected on the bacteria after staining with sera were selected,defined and measured using the functions Image>Adjust>Threshold andAnalyze>Measure of ImageJ. *=P<0.05; **=P<0.01; ***=P<0.001.

FIGS. 10A and 10B: Opsonophagocytic activity of antibodies in mice sera.(FIG. 10A) RAW mouse macrophages were incubated with Streptococcuspneumoniae type 1, pre-incubated with mouse sera from the adjuvant orMP+adjuvant groups. Indicated are the ratios of adherence of bacteria tocells, or uptake of bacteria inside cells. Data are represented asmeans+/−SEM of three independent experiments. *=P<0.05. (FIG. 10B) RAWmouse macrophages have been incubated with Streptococcus pneumoniae type1, pre-incubated with mouse sera from the adjuvant or MP+adjuvantgroups. Indicated is the percentage of taken up bacteria survivinginside the cells after 1 hour. Data are represented as means+/−SEM ofthree independent experiments. *=P<0.05.

FIGS. 11A through 11E: Intranasal immunization of C57BL/6 mice with MPof S. pneumoniae increases survival against intranasal infection withserotype 3. In particular, immunization with MPs of serotype 3 confers100% protection against serotype 3 infection in comparison to PCV13immunization, which confers 75% protection. B-cell deficient miceimmunized with MPs of serotype 3 are not protected against intranasalinfection with serotype 3, strongly suggesting that protection providedby MPs is antibody-dependent. (FIG. 11A) Percentage of mice thatsurvived after pneumococcal challenge. 10 mice in the group of “BCell—(deficient mice)”, 10 mice in the group “Adjuvant”, 20 mice per allother groups. *=P<0.05; ****=P<0.0001. (FIG. 11B) CFUs in lungs of miceafter sacrifice. Each dot represents one mouse. ***=P<0.001. (FIGS.11C-11E) CFUs in blood of mice. Each line represents one mouse.

SUMMARY OF THE INVENTION

The present invention relates to the following items. The subject matterdisclosed in the items below should be regarded disclosed in the samemanner as if the subject matter were disclosed in patent claims.

-   -   1. An isolated Streptococcus pneumoniae membrane vesicle        microparticle (MP), wherein said MP comprises:        -   i. the protein Ply at the level of ≥0.070 μg/μg total            protein in the MP;        -   ii. the protein LytA at the level of ≥0.070 μg/μg total            protein in the MP;        -   iii. the protein PspC at the level of ≥0.130 μg/μg total            protein in the MP; or        -   iv. the protein RrgB at the level of ≥0.020 μg/μg total            protein in the MP.    -   2. The microparticle according to item 1, further comprising a        capsular polysaccharide of a capsular serotype of Streptococcus        pneumoniae, preferably at a level of ≥0.001, more preferably        ≥0.01, most preferably ≥0.1 μg/μg total protein in the MP.    -   3. The microparticle according to any of the preceding items,        comprising the protein Ply at the level of ≥0.070 μg/μg total        protein in the MP.    -   4. The microparticle according to any of the preceding items,        comprising the protein Ply at the level of ≥0.15, preferably        ≥0.2, more preferably ≥0.3, most preferably ≥0.35 μg/μg total        protein in the MP.    -   5. The microparticle according to any of the preceding items,        comprising the protein LytA at the level of ≥0.070 μg/μg total        protein in the MP.    -   6. The microparticle according to any of the preceding items,        comprising the protein LytA at the level of ≥0.08, preferably        ≥0.09, yet more preferably ≥0.10, most preferably ≥0.20 μg/μg        total protein in the MP.    -   7. The microparticle according to any of the preceding items,        comprising the protein PspC at the level of ≥0.130 μg/μg total        protein in the MP.    -   8. The microparticle according to any of the preceding items,        comprising the protein PspC at the level of ≥0.15, preferably        ≥0.18, more preferably ≥0.20, most preferably ≥0.3 μg/μg total        protein in the MP.    -   9. The microparticle according to any of the preceding items,        comprising the protein RrgB at the level of ≥0.020 μg/μg total        protein in the MP.    -   10. The microparticle according to any of the preceding items,        comprising the protein RgrB at the level of ≥0.022, preferably        ≥0.025, most preferably ≥0.028 μg/μg total protein in the MP.    -   11. The microparticle according to any of the preceding items,        further comprising the protein PhtD.    -   12. The microparticle according to any of the preceding items,        comprising the protein PhtD at a level being at least 2-fold,        more preferably at least 2.5-fold compared to Streptococcus        pneumoniae membrane vesicles obtained in liquid culture in terms        of μg/μg total protein in the particle or the vesicle,        respectively.    -   13. The microparticle according to any of the preceding items,        further comprising the protein RrgA.    -   14. The microparticle according to any of the preceding items,        comprising the protein RrgA at the level of ≥0.02, preferably        ≥0.05, more preferably ≥0.10, most preferably ≥0.2 μg/μg total        protein in the MP.    -   15. The microparticle according to any of the preceding items,        comprising the protein IgA.    -   16. The microparticle according to any of the preceding items,        comprising the protein IgA at the level of ≥0.02, preferably        ≥0.05, more preferably ≥0.10, most preferably ≥0.2 μg/μg total        protein in the MP.    -   17. The microparticle according to any of the preceding items,        wherein the protein Ply comprises a sequence having at least        80%, preferably 85%, more preferably 90%, yet more preferably        95%, most preferably 100% sequence identity to SEQ ID NO: 1.    -   18. The microparticle according to any of the preceding items,        wherein the protein LytA comprises a sequence having at least        80%, preferably 85%, more preferably 90%, yet more preferably        95%, most preferably 100% sequence identity to SEQ ID NO: 2.    -   19. The microparticle according to any of the preceding items,        wherein the protein PspC comprises a sequence having at least        40%, preferably 80%, more preferably 85%, yet more preferably        90%, still more preferably 95%, most preferably 100% sequence        identity to SEQ ID NOs: 3.    -   20. The microparticle according to any of the preceding items,        wherein the protein PspC comprises a sequence having at least        80%, preferably 85%, more preferably 90%, yet more preferably        95%, most preferably 100% sequence identity to any one of SEQ ID        NOs: 3-14.    -   21. The microparticle according to any of the preceding items,        wherein the protein RgrB comprises a sequence having at least        80%, preferably 85%, more preferably 90%, yet more preferably        95%, most preferably 100% sequence identity to SEQ ID NO: 15.    -   22. The microparticle according to any of the preceding items,        wherein the protein PhtD comprises a sequence having at least        80%, preferably 85%, more preferably 90%, yet more preferably        95%, most preferably 100% sequence identity to SEQ ID NO: 16.    -   23. The microparticle according to any of the preceding items,        comprising the protein RgrA, wherein the protein RgrA comprises        a sequence having at least 80%, preferably 85%, more preferably        90%, yet more preferably 95%, most preferably 100% sequence        identity to SEQ ID NO: 17.    -   24. The microparticle according to any of the preceding items,        comprising the protein IgA, wherein the protein IgA comprises a        sequence having at least 80%, preferably 85%, more preferably        90%, yet more preferably 95%, most preferably 100% sequence        identity to SEQ ID NO: 18.    -   25. The microparticle according to any of the preceding items,        wherein the MP is 5-300 nm in diameter, preferably 15-175 nm.    -   26. The microparticle according to any of the preceding items,        wherein the MP is 10-125 nm in diameter.    -   27. The microparticle according to any of the preceding items,        wherein the MP is derived from a Streptococcus pneumoniae strain        selected from any serotype 1 strain, any serotype 3 strain,        TIGR4, P1031 and A66, preferably TIGR4.    -   28. The microparticle according to any of the preceding items,        obtainable by:        -   a. Culturing host cells of a Streptococcus pneumoniae strain            on blood agar plates;        -   b. harvesting the cultured host cells;        -   c. centrifuging the harvested host cells at 17,000 g for 30            minutes at +4 C;        -   d. subjecting the supernatant to filtration through a 0.22            μm filter;        -   e. centrifuging the filtered supernatant at 120,000 g for 2            h at +4 C;        -   f. washing the pellets from e) twice in phosphate-buffered            saline with sedimentations at 120,000 g for 2 h at +4 C;        -   g. resuspending the pellet in 1 ml phosphate-buffered            saline;        -   h. adjusting the resuspended pellet to 50% (w/v) Optiprep™            density gradient medium in a total volume of 2 ml and            overlaying with one fraction of 30% (w/v) Optiprep™ (9 ml)            followed by a fraction of 5% (w/v) Optiprep™ (3 ml);        -   i. centrifuging the gradients at 250,000×g for 3 hours at 4°            C.;        -   j. collecting the 4 ml top fraction, containing the membrane            microparticle; and        -   k. washing the microparticle three times phosphate-buffered            saline with sedimentations of 250,000×g for 2 hours at +4°            C.    -   29. A composition comprising a microparticle according to any of        items 1-28.    -   30. The composition according to item 29, wherein the        composition is devoid of whole Streptococcus pneumoniae cells.    -   31. The composition according to any of items 29-30, further        comprising capsular polysaccharides from Streptococcus        pneumoniae.    -   32. The composition according to any of items 29-31, comprising        MPs in an amount of 1 μg/ml, preferably 5 μg/ml, more preferably        10 μg/ml, most preferably 100 μg/ml.    -   33. The composition according to any of items 29-32, further        comprising an adjuvant.    -   34. The composition according to any of items 29-33, further        comprising an adjuvant comprising aluminium hydroxide.    -   35. The composition according to any of items 29-34, being an        immunogenic composition.    -   36. The composition according to any of items 29-35, being an        immunogenic composition capable of eliciting opsonophagocitic        antibodies against Streptococcus pneumoniae when administered to        a mammalian host.    -   37. The composition according to any of items 29-36, being an        immunogenic composition is capable of eliciting serotype        independent antibodies against Streptococcus pneumoniae when        administered to a mammalian host.    -   38. The composition according to any of items 29-37, being an        immunogenic composition is capable of eliciting antibodies        against Streptococcus pneumoniae serotype 3 when administered to        a mammalian host.    -   39. The composition according to any of items 29-38, further        comprising capsular polysaccharides of a capsular serotype of        Streptococcus pneumoniae.    -   40. The composition according to any of items 29-39, being a        vaccine.    -   41. The composition according to any of items 29-40, for use in        a method for inducing protective immunity against Streptococcus        pneumoniae in a subject.    -   42. The composition according to item 41, for use according to        item 41, wherein the immunity is protective against a condition        selected from pneumococcal sinusitis, pneumococcal otitis,        pneumococcal pneumonia and invasive pneumococcal disease        including but not limited to pneumococcal sepsis and        pneumococcal meningitis, preferably invasive pneumococcal        disease.    -   43. A use of a microparticle according to any of items 1-28 in        an immunogenic composition.    -   44. The use according to item 43, wherein the immunogenic        composition is a vaccine.    -   45. A use of a microparticle according to any of items 1-28 in        the manufacture of a vaccine.    -   46. A method for manufacturing a vaccine, comprising:        -   a. Providing a microparticle according to any of items 1-28;        -   b. Providing an adjuvant;        -   c. Mixing the microparticle and the adjuvant in a suitable            vehicle in order to produce a vaccine.    -   47. A method for producing an isolated Streptococcus pneumoniae        microparticle according to any of items 1-28, comprising:        -   a. providing a Streptococcus pneumoniae host cell;        -   b. culturing said host cell under conditions allowing the            production of a microparticle according to any of items 1-28            by the host cell; and        -   c. isolating the microparticle thus produced.    -   48. The method according to item 47, wherein the host cell is of        a strain selected from any serotype 1 strain, any serotype 3        strain, TIGR4, P1031 and A66, preferably TIGR4.    -   49. The method according to any of items 47-48, wherein the        culturing is carried out in solid phase.    -   50. The method according to any of items 47-49, wherein the        culturing is carried out in solid phase on blood agar.    -   51. The method according to any of items 47-50, wherein        isolating the microparticle comprises density gradient        centrifugation.

DETAILED DESCRIPTION

The inventors isolated membrane vesicles (MV_(L)) from S. pneumoniae ofserotype 4 grown in liquid cultures as previously described in theliterature (1). They also isolated a novel type of membrane particlestermed microparticles (MP) using a method involving growth of thebacteria overnight on blood agar plates. FIG. 1 shows the appearance ofthe MV_(L) and MP using electron microscopy and atomic force microscopyas well as their size distribution. Microparticles from S. pneumoniaeTIGR4 grown on plates (MP) were smaller in size on average than MV_(L)coming from liquid media (MV_(L)). However, MP from S. pneumoniaeserotype 3 display a different size distribution, containing less of thesmallest particles and more of the largest ones.

Biochemical comparison of MV_(L) and MP preparations from TIGR4 revealeddifferent properties for the MP and MV_(L). MP isolated from platescarried certain pneumococcal proteins to a higher extent than liquidgrown MV_(L). They were particularly enriched in pneumolysin, the majorpore-forming toxin in S. pneumoniae, as shown in FIG. 2D.

The novel MP were characterized using in vitro cell assays, and it wasshown that consistent with the quantitated levels, the MP contain morebiological pneumolysin activity than the MV_(L) (FIG. 3 ). It wasfurther shown that the MP are taken up by epithelial cells (FIG. 4A-D)but are not cytotoxic (FIG. 4E). Using human monocyte-derived dendriticcells it was shown that both MP and MV_(L) are internalized into DCs,induce maturation of DCs (FIG. 5 ), and elicit differential cytokineresponses (FIG. 6 ).

Next, the inventors investigated the potential of MP to protect miceusing experimental models of pneumonia and invasive disease (FIGS. 7-11). The results showed that 80% of mice (wild type C57BL/6 mice)immunized with MP from serotype 4 (combined with aluminium hydroxide asadjuvant) survived after an intranasal pneumococcal infection caused byserotype 1 (heterologous challenge, cross protection). ELISA analysisshowed that mice immunized with MP had a significantly higherMP-specific IgG response in comparison to control mice (treated onlywith the adjuvant) (FIG. 8 ). Immunization with serotype 3 and thenchallenge with serotype 3 led to a 100% protection (FIG. 11 ).Immunization with serotype 4 MP and then challenge with serotype 3 (aserotype that has been problematic in current vaccines) gave aprotection of ca 50% (FIG. 11 ). Furthermore, immunofluorescencestainings showed that sera from mice immunized with MP from serotype 4contained antibodies specific against multiple other serotypes, such asserotypes 1, 3 and 6B (FIG. 9 ).

In conclusion, it was found that MP protect mice against pneumococcalinfection, and, most importantly, such protection isserotype-independent to a significant degree, in contrast to theprotection conferred by currently available vaccines (polysaccharide orconjugated vaccines) that are, as mentioned in the background section,strictly serotype-dependent. The MP immunisation furthermore elicitedprotection against serotype 3 which is a challenge for current vaccines.Indeed, immunization with the currently clinically used vaccine PCV13provided significantly less protection to mice than MP isolated fromserotype 3 (FIG. 11 ).

Moreover, for protein based vaccines that are under development today,there is a problem in obtaining opsonophagocytic capability.Importantly, the present data show that using MP as a vaccine affectsopsonophagocytosis (Example 3, FIG. 10 ). Last but not least, availablepneumococcal vaccines (PCVs) are extremely important to fightpneumococcal diseases in developing countries. However, the costs toproduce current vaccines are very high. An important benefit with thepresent approach is a drop in production costs since isolation ofmicroparticles does not require high-cost preparations withconjugations, instead the method of the present invention is more simplebased on bacterial growth combined with high speed-centrifugation.

Isolated Streptococcus pneumoniae Membrane Vesicles and Microparticles

As discussed in the Background section, a few types of Streptococcuspneumoniae membrane vesicles have been obtained in liquid culture (1).The solid phase culture method used by the inventors resulted in a noveltype of particles termed microparticles having different andadvantageous properties as discussed above and shown in the Examples. Asshown in Table 1, the microparticles differ from the known membranevesicles, MV_(L) in terms of relative protein expression.

TABLE 1 Quantification of proteins present in membrane vesicles, MV_(L),from serotype 4 strain TIGR4 and microparticles, MP, from serotype 4 and3. Percentage sequence identities of protein homologues in referencestrains for serotype 1 (P1031) and serotype 3 (A66) are shown. Theamounts are indicated in μg/μg total protein or relative amount inpercentage. MP Homology in Homology in Protein MV_(L) (TIGR4) MP (Type3) P1031* A66** Ply 0.055 μg 0.406 μg 0.314 μg 99% 99% LytA 0.050 μg0.100 μg 0.266 μg 99% 100%  PspC 0.118 μg 0.300 μg Not determined* 85%(PspC 45% (PspC (PspC3.4) 2.1) 11.4) RrgB 0.016 μg 0.028 μg AbsentAbsent Absent PhtD 100% 266% 5793% 95% 87% *Because of the high geneticdiversity of the pspC gene among pneumococcal serotypes it was notpossible to use the anti TIGR4 PspC antibody (which was specific for theTIGR4 strain) to detect the serotype 3 PspC

Thus, in a first aspect of the present invention, there is providedisolated Streptococcus pneumoniae microparticle (MP) (a novel type ofpneumococcal membrane vesicle particle), wherein said MP comprise eitherone, two, three or four of the following proteins at the designatedlevels:

-   -   i. the protein Ply at the level of ≥0.070 μg/μg total protein in        the MP;    -   ii. the protein LytA at the level of ≥0.070 μg/μg total protein        in the MP;    -   iii. the protein PspC at the level of ≥0.130 μg/μg total protein        in the MP; and/or    -   iv. the protein RrgB at the level of ≥0.020 μg/μg total protein        in the MP.

The microparticles may further comprise capsular polysaccharides of acapsular serotype of Streptococcus pneumoniae, preferably at a level of≥0.001, more preferably ≥0.01, most preferably ≥0.1 μg/μg total proteinin the MP.

The microparticle may comprise the protein Ply at the level of ≥0.070μg/μg total protein in the MP. The microparticle may comprise theprotein Ply at the level of ≥0.15, preferably ≥0.2, more preferably≥0.3, most preferably ≥0.35 μg/μg total protein in the MP.

The microparticle may comprise the protein LytA at the level of ≥0.070μg/μg total protein in the MP. The microparticle may comprise theprotein LytA at the level of ≥0.08, preferably ≥0.09, yet morepreferably ≥0.1 μg/μg, most preferably ≥0.2 μg/μg total protein in theMP.

The microparticle, MP, may comprise the protein PspC at the level of≥0.130 μg/μg total protein in the MP. The microparticle may comprisecomprising the protein PspC at the level of ≥0.15, preferably ≥0.18,more preferably ≥0.2, most preferably ≥0.3 μg/μg total protein in theMP.

The microparticle may comprise the protein RrgB at the level of ≥0.02μg/μg total protein in the MP. The microparticle may comprise theprotein RgrB at the level of ≥0.022, preferably ≥0.025, most preferably≥0.028 μg/μg total protein in the MP.

The microparticle may further comprise the protein PhtD. The proteinPhtD may be present at a level being at least 2-fold, more preferably atleast 2.5-fold compared to Streptococcus pneumoniae membrane vesiclesobtained in liquid culture in terms of μg/μg total protein in theparticle or the vesicle, respectively.

The microparticle may further comprise the protein RrgA. The proteinRrgA may be present at the level of ≥0.02, preferably ≥0.05, morepreferably ≥0.10, most preferably ≥0.2 μg/μg total protein in the MP.

The microparticle may further comprise the Streptococcus pneumoniaeprotein IgA. The protein IgA may be present at the level of ≥0.02,preferably ≥0.05, more preferably ≥0.10, most preferably ≥0.2 μg/μgtotal protein in the MP.

The protein Ply may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to SEQ ID NO: 1.

The protein LytA may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to SEQ ID NO: 2.

The protein PspC may comprise a sequence having at least 40% or 70%,preferably 80%, more preferably 85%, yet more preferably 90%, still morepreferably 95%, most preferably 100% sequence identity to SEQ ID NOs: 3.

The protein PspC may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to any one of SEQ ID NOs:3-14.

The protein RgrB may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to SEQ ID NO: 15.

The protein PhtD may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to SEQ ID NO: 16.

The protein RgrA may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to SEQ ID NO: 17.

The protein IgA may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to SEQ ID NO: 18.

Thus, the first aspect encompasses (but is not limited to) embodimentsdisclosed in Table 2 below.

TABLE 2 Minimum levels of designated proteins in various embodiments ofthe first aspect. A blank cell indicates that the protein is optionali.e. may be absent or present at any level. Units: μg/μg total protein,except for PhtD, where the amount is in comparing to Streptococcuspneumoniae membrane vesicles obtained in liquid culture in terms ofμg/μg total protein in the particle or the vesicle, respectively.Embodiment# Ply LytA PspC PhtD RgrB 1 0.070 2 0.070 0.070 3 0.070 0.1304 0.070 2-FOLD 5 0.070 0.070 0.130 6 0.070 0.070 2-FOLD 7 0.070 0.0700.130 2-FOLD 8 0.070 0.130 2-FOLD 9 0.070 10 0.130 11 2-FOLD 12 0.0700.130 13 0.070 2-FOLD 14 0.070 0.130 2-FOLD 15 0.130 2-FOLD 16 0.0700.020 17 0.070 0.070 0.020 18 0.070 0.130 0.020 19 0.070 2-FOLD 0.020 200.070 0.070 0.130 0.020 21 0.070 0.070 2-FOLD 0.020 22 0.070 0.070 0.1302-FOLD 0.020 23 0.070 0.130 2-FOLD 0.020 24 0.070 0.020 25 0.130 0.02026 2-FOLD 0.020 27 0.070 0.130 0.020 28 0.070 2-FOLD 0.020 29 0.0700.130 2-FOLD 0.020 30 0.130 2-FOLD 0.020 31 0.070 32 0.070 0.070 330.070 0.130 34 0.070 2-FOLD 35 0.070 0.070 0.130 36 0.070 0.070 2-FOLD37 0.070 0.070 0.130 2-FOLD 38 0.070 0.130 2-FOLD 39 0.070 40 0.130 412-FOLD 42 0.070 0.130 43 0.070 2-FOLD 44 0.070 0.130 2-FOLD 45 0.1302-FOLD 46 0.070 0.020 47 0.070 0.070 0.020 48 0.070 0.130 0.020 49 0.0702-FOLD 0.020 50 0.070 0.070 0.130 0.020 51 0.070 0.070 2-FOLD 0.020 520.070 0.070 0.130 2-FOLD 0.020 53 0.070 0.130 2-FOLD 0.020 54 0.0700.020 55 0.130 0.020 56 2-FOLD 0.020 57 0.070 0.130 0.020 58 0.0702-FOLD 0.020 59 0.070 0.130 2-FOLD 0.020 60 0.130 2-FOLD 0.020

The microparticles may be 5-300 nm in diameter, preferably 10-125 nm indiameter.

The microparticles may be derived from any Streptococcus pneumoniaestrain, but is preferably selected from a group consisting of allserotype 3 strains, all serotype 1 strains, TIGR4, P1031 and A66, mostpreferably TIGR4.

The microparticles according to the first aspect may be obtainable by:

-   -   a. culturing host cells of a Streptococcus pneumoniae strain on        blood agar plates or other plates;    -   b. harvesting the cultured host cells;    -   c. centrifuging the harvested host cells at 17,000 g for 30        minutes at +4° C.;    -   d. subjecting the supernatant to filtration through a 0.22 μm        filter;    -   e. centrifuging the filtered supernatant at 120,000 g for 2 h at        +4° C.;    -   f. washing the pellets from e) twice in phosphate-buffered        saline with sedimentations at 120,000 g for 2 h at +4° C.;    -   g. resuspending the pellet in 1 ml phosphate-buffered saline;    -   h. adjusting the resuspended pellet to 50% (w/v) Optiprep™        density gradient medium in a total volume of 2 ml and overlaying        with one fraction of 30% (w/v) Optiprep™ (9 ml) followed by a        fraction of 5% (w/v) Optiprep™ (3 ml);    -   i. centrifuging the gradients at 250,000×g for 3 hours at 4° C.;    -   j. collecting the 4 ml top fraction, containing the membrane        microparticles;    -   k. washing the microparticles three times phosphate-buffered        saline with sedimentations of 250,000×g for 2 hours at +4° C.

Compositions and Uses

In a second aspect of the present invention, there is provided acomposition comprising a microparticle according to the first aspect.The composition may be devoid of whole Streptococcus pneumoniae cells.The composition may optionally comprise capsular polysaccharides fromStreptococcus pneumoniae.

The composition according to the second aspect may comprise MP in anamount of 1 μg/ml, preferably 5 μg/ml, more preferably 10 μg/ml, mostpreferably 100 μg/ml (the concentrations refer to the total proteincontent of MP per ml).

The composition may further comprise an adjuvant. The adjuvantpreferably comprises aluminium hydroxide.

The composition may be an immunogenic composition. Preferably, theimmunogenic composition is capable of eliciting opsonophagociticantibodies against Streptococcus pneumoniae when administered to amammalian host. Preferably, the immunogenic composition is capable ofeliciting serotype independent antibodies against Streptococcuspneumoniae when administered to a mammalian host. Preferably, theimmunogenic composition is capable of eliciting antibodies againstStreptococcus pneumoniae serotype 3 when administered to a mammalianhost.

The composition may be formulated as a vaccine.

In a third aspect of the present invention, there is provided acomposition according to the second aspect, for use in a method forinducing protective immunity against Streptococcus pneumoniae in asubject. The third aspect also encompasses a method for inducingprotective immunity against Streptococcus pneumoniae in a subject inneed thereof, comprising administering an effective amount of thecomposition according to the second aspect to the subject. The thirdaspect also encompasses the use of a composition according to the secondaspect in the manufacture of a vaccine for immunization againstStreptococcus pneumoniae.

The protective immunity may be an immunity reducing the likelihood of acondition selected from pneumococcal sinusitis, pneumococcal otitis,pneumococcal pneumonia and invasive pneumococcal disease including butnot limited to pneumococcal sepsis and pneumococcal meningitis,preferably invasive pneumococcal disease. Preferably, the subject to beimmunized is a young child (e.g. less than 7 years of age) or an elderlyperson (e.g. over 65 years of age), but also other age groups could betargeted.

For immunization, the composition may be administered to the subject invarious manners known in the art, including but not limited to by way ofinjection (e.g. intramuscular, intracutaneous, subcutaneous,intravenous), buccal, oral and intranasal administration as well asinhalation.

In a fourth aspect there is provided a use of a microparticle accordingto the first aspect in an immunogenic composition. The immunogeniccomposition may be a vaccine.

In a fifth aspect, there is provided a use of a microparticle accordingto the first aspect in the manufacture of a vaccine.

In a sixth aspect, there is provided a method for manufacturing avaccine, comprising:

-   -   a. Providing a microparticle according to the first aspect    -   b. Providing an adjuvant, such as aluminium hydroxide;    -   c. Mixing the microparticle and the adjuvant in a suitable        vehicle in order to produce a vaccine.

Methods for Producing Streptococcus pneumoniae Microparticles

In a sixth aspect of the present invention there is provided a methodfor producing an isolated Streptococcus pneumoniae microparticleaccording to the first aspect, comprising:

-   -   a. providing a Streptococcus pneumoniae bacterial cell;    -   b. culturing said bacterial cell under conditions allowing the        production of a microparticle according to the first aspect by        the bacterial cell; and    -   c. isolating the microparticle thus produced.

The bacterial cell may be from any pneumococcal strain, but ispreferably TIGR4, a serotype 1 and/or a serotype 3 strain.

The culturing may be carried out using plates or liquid culture.Preferably, the culturing is carried out in solid phase, most preferablyon blood agar or other plates. Isolating the microparticles may comprisea density gradient centrifugation step.

General Aspects Relating to the Present Disclosure

The term “comprising” is to be interpreted as including, but not beinglimited to. All references are hereby incorporated by reference. Thearrangement of the present disclosure into sections with headings andsubheadings is merely to improve legibility and is not to be interpretedlimiting in any way, in particular, the division does not in any waypreclude or limit combining features under different headings andsubheadings with each other.

EXAMPLES

The following examples are not to be regarded as limiting. For furtherinformation on the experimental details, the skilled reader is directedto a separate section titled Materials and Methods.

Example 1: Isolation and In Vitro Characterization of Novel PneumococcalMicroparticles

S. pneumoniae TIGR4 Produces Microparticles that Differ Depending onBacterial Growth Conditions.

Membrane vesicles were isolated and purified from Streptococcuspneumoniae serotype 4 strain TIGR4 grown in liquid medium (MV_(L)).Microparticles (MP) were isolated from pneumococci grown on blood agarplates. Preparations were analyzed by transmission electron microscopy(TEM) and atomic force microscopy, and revealed spherical bodiessurrounded by membranous structures (FIG. 1A). The distribution of thesize varied between MV_(L), MP from TIGR4 and MP from serotype 3 (FIG.1B). In total 503 MV_(L), 378 MP from TIGR4 and 523 MP from serotype 3were analyzed. As a control we also studied an isogenic pneumolysinmutant strain (TIGR4Δply) and found a similar size distribution. Toreveal shedding of MV_(L) or MP from the surface of bacteria by TEM wethen used an isogenic unencapsulated mutant in TIGR4 (T4R) grown inliquid culture. In the absence of the thick bacterial capsule, MV_(L)were clearly visible, emanating from the plasma membrane of S.pneumoniae into the extracellular milieu (FIG. 1C-D).

Proteomic Analysis Revealed Differences in Protein Content BetweenMV_(L) from Liquid Culture or MP from Plates

To analyze the protein content of both pneumococcal preparations,purified MV_(L) and MP were subjected to SDS-PAGE analysis. Distinctprotein patterns in MV_(L) and MPs as compared with proteins present inwhole cell lysates or bacterial supernatants (FIG. 2A), were identified.Interestingly, the proteomic content in MV_(L) and MP fractionsdiffered, suggesting a differential enrichment of proteins in thedifferent types of particles. We further conducted proteomic analysisusing tandem mass spectrometry. In total, in TIGR4, we identified 317proteins in MV_(L) (Table 3), and 216 in MP (Table 4), of which 184proteins were found in both preparations. 133 proteins were unique toMV_(L) and 32 to MP (FIG. 2B). In serotype 3 we identified 462 proteinsin MV_(L) (Table 5) and 344 in MP (Table 6). All detected proteins werefurther classified according to their subcellular localization based onthe combination of LocateP and GO annotation (FIG. 2C) (11). A majorityof the proteins identified in MV_(L) and MP were cytosolic proteins,followed by multi-transmembrane proteins, lipoproteins, N-terminallyanchored proteins, secretory proteins and cell wall-associated proteins.While MV_(L) appeared to be more enriched in cytosolic protein and cellwall-associated proteins than MP from TIGR4, the latter harbored ahigher percentage of multi-transmembrane proteins, lipoproteins,N-terminally anchored proteins and secretory proteins.

We further analyzed the mass spectrometry data in order to determine thepresence of 30 known pneumococcal virulence factors in the particles.Both types of particles contained approximately half of these virulencefactors present in TIGR4. Just two were unique to MV_(L), CbpD (12) andBgaA (13), and one to MP, IgA (10).

Based on the proteomic results, we next performed immunoblot analysis onspecific pneumococcal proteins and virulence factors (FIG. 2D). Amongcytosolic proteins we investigated the presence of the cytotoxinpneumolysin (Ply), which is a pore-forming toxin and one of the mostimportant virulence factors in pneumococcal pathogenesis (2), GAPDH as acytosolic marker (14), and LytA, the major autolysin of Streptococcuspneumoniae (3). The most striking observation was the high enrichment ofPly especially in MP, which also contained more LytA than MV_(L).Smaller differences were also observed in the enrichment of virulencefactors belonging to other subcellular localizations. While MV_(L)displayed a higher amount of Pneumococcal surface antigen A (PsaA) (15)and Sortase A (SrtA) (16) than MP, the latter contained morePneumococcal surface protein C (PspC) (17) and Polyhistidine triadprotein D (PhtD) (9) (FIG. 2D). The major component of the pneumococcalpilus, RrgB (6), was equally found in both particle preparations.

These findings suggest the presence of a still uncharacterized mechanismfor the selective secretion and enrichment of specific proteins andvirulence factors of Streptococcus pneumoniae in these particlepreparations.

A Biologically Active Pneumolysin was Found in Membrane Microparticles

The enrichment of Ply in both MV_(L) and MP prompted us to determinewhether particle-associated Ply was functional and able to lyseerythrocytes in a hemolytic assay. For both types of particles weobserved a dose dependent hemolysis (FIG. 3 ), however, Ply from MPshowed higher activity than from MV_(L), which is consistent with theobservation that MP contain more Ply. Notably, hemolysis seemed to beentirely caused by Ply, and particles prepared from a TIGR4 mutantlacking Ply (TIGR4Δply) exhibited no hemolytic activity.

MP are Taken Up by A549 Epithelial Cells, but are not Cytotoxic.

MV in other Gram-positive bacteria have been described to function asvehicle to deliver vesicle-associated proteins to human cells (18) (19).To test whether pneumococcal MP can deliver pneumococcal proteins, weincubated MP from TIGR4 with A549 lung epithelial cells for 24 hours andvisualized Ply and LytA by immunofluorescence staining (FIG. 4A-C). Byanalyzing orthogonal views of A549 cells treated with MP, we showed byimmunofluorescence that MP can be taken up, with Ply and LytA detectableinside the cells (FIG. 4B-C).

To confirm our observation we further analyzed lysates of A549 cellstreated with MP by immunoblotting with anti-Ply antibody (FIG. 4D). Inorder to minimize extracellular associated particles, treated cells wereexcessively washed before lysates were made. Consistent with ourimmunofluorescence results, western blotting revealed a dose dependentincrease in Ply further suggesting that MP can be internalized.

Next we asked whether proteins associated with MV_(L) or MP, and inparticular Ply, may contribute to cytotoxic effects on cultured humancells. To assess toxicity, A549 cells were treated with differentconcentrations of MV_(L) and MP for 24 hours, stained with fixableviability dye and analyzed by flow cytometry (FIG. 4E). The differencein cell death between samples incubated with MV_(L) or MPs and thenegative control sample was not statistically significant, suggestingthat Ply is not cytotoxic to the cells at concentrations present inMV_(L) or MP.

MV_(L) and MP are Internalized by Human Monocyte-Derived Dendritic Cells(DCs) and Induce their Maturation.

Since we found that MP can be internalized by epithelial cells, we thenstudied potential interactions of MV_(L) or MP with innate immune cells,focusing on DCs. First we assessed cytotoxicity by incubating DCs withMV_(L) or MP, stained with fixable viability dye and Annexin V, and thenanalyzed using flow cytometry (FIG. 5A). No significant increase incytotoxicity was detected in samples incubated with MP, while additionof MV_(L) displayed cytotoxic effects at the two highest concentrationsused. While both preparations were considerably less cytotoxic than livebacteria, with 70-80% cytotoxic cells after 24 hours of incubation, morestudies are needed to assess the mechanism behind the increasedcytotoxicity of MV_(L) from liquid cultures. Moreover, the percentage ofapoptotic cells did not change after incubation with either of theparticle preparations.

Then we investigated uptake of MP by DCs. First we incubated DCs with MPfor 1 hour, lysed the cells and performed SDS-PAGE and immunoblotting oncell lysates to check for presence of Ply. Indeed, Ply was found in thecell lysates (FIG. 5B), suggesting that DCs internalize MP. Next, DCswere challenged for 30 minutes with inhibitors of actin polymerizationand phosphatidylinositol 3-kinase (PI3K), cytochalasin and wortmannin,to block active phagocytosis, or with an inhibitor of lipid rafts,methyl-β-cyclodextrin, to block fusion of membranes. Subsequently, cellswere incubated with MP for 1 hour, lysed, and the same protocol forimmunoblotting was followed. Interestingly, blocking active phagocytosisor membrane fusion resulted in less Ply. However, when all inhibitorswere used together, Ply still remained present in the lysates,suggesting either that the particles adhered to the cells or wereinternalized by another mechanism, not investigated in this study.

Taken together these data suggest that MP can function as a vehicle todeliver bacterial components to host cells. Since DCs are professionalantigen presenting cells, we then assessed whether MV_(L) or MP couldinfluence DC maturation. Cells were incubated for 24 hours with MV_(L)or MP, and then stained for MHCII and the co-stimulatory molecule CD86,and analyzed by flow cytometry (FIG. 5C). A dose-dependent increase inthe presence of both markers on the surface of DCs was detected whencells were incubated with the particles. MV_(L) appeared to induce morematuration than MP, and similar levels of maturation were obtained withLPS.

MV_(L) and MP Induce Pro-Inflammatory Cytokine Responses in DCs

Since OMVs from Gram-negative have been shown to trigger a potent innateimmune response (20) (21), we studied whether also pneumococcal MV_(L)or MP could affect pro-inflammatory cytokine responses. DCs we incubatedwith MV_(L) or MP for 24 hours, and ELISA assays were used onsupernatants to determine concentrations of IL-6 (FIG. 6A), IL-8 (FIG.6B), IL-10 (FIG. 6C) and TNF (FIG. 6D). MP induced release of IL-8 andlow levels of TNF, while no induction was seen for IL-6 and IL-10. Incontrast, MV_(L) induced all four cytokines tested to levels higher thanlive bacteria. None of the particles induced IL-1β or IL-12 (data notshown).

TABLE 3 Proteins identified in MV_(L) from TIGR4. The accession numbersrefer to SwissProt on the date of filing. # Accession Coverage Peptides#AAs Score Description Localization Q97RH0 76.86 47 350 2615.53Lipoprotein OS = Streptococcus Lipid pneumoniae serotype 4 (strain ATCCanchored BAA-334/TIGR4) GN = SP_0845 PE = 4 SV = 1 − [Q97RH0_STRPN]Q2MGF6 82.45 59 490 1620.74 Lysozyme OS = Streptococcus Secretorypneumoniae serotype 4 (strain ATCC (released) BAA-334/TIGR4) GN = lytCPE = 4 (with CS) SV = 1 − [Q2MGF6_STRPN] Q97NK0 73.39 64 883 1364.43Aldehyde-alcohol dehydrogenase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_2026 PE = 3SV = 1 − [Q97NK0_STRPN] P18791 72.99 48 659 1307.17 Oligopeptide-bindingprotein AmiA Lipid OS = Streptococcus pneumoniae anchored serotype 4(strain ATCC BAA-334/ TIGR4) GN = amiA PE = 1 SV = 3 − [AMIA_STRPN]Q9L7Q2 55.82 91 1906 1166.89 Zinc metalloprotease ZmpB Multi- OS =Streptococcus pneumoniae trans- serotype 4 (strain ATCC BAA-334/membrane TIGR4) GN = zmpB PE = 3 SV = 2 − [ZMPB_STRPN] Q97T80 58.3 871856 1012.57 Zinc metalloprotease ZmpC LP × TG OS = Streptococcuspneumoniae Cell-wall serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN= zmpC PE = 3 SV = 1 − [ZMPC_STRPN] Q97NQ8 59.59 75 1225 936.07DNA-directed RNA polymerase subunit Intracellular beta′ OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rpoC PE = 3 SV = 1 − [RPOC_STRPN] P59213 74.47 48 423 838.2Maltose/maltodextrin-binding protein Lipid OS = Streptococcus pneumoniaeanchored serotype 4 (strain ATCC BAA-334/ TIGR4) GN = malX PE = 1 SV = 1− [MALX_STRPN] P64022 75.61 46 693 803.77 Elongation factor G OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = fusA PE = 3 SV = 1 − [EFG_STRPN] P0C2J9 92.99 46 471753.38 Pneumolysin OS = Streptococcus Intracellular pneumoniae serotype4 (strain ATCC BAA-334/TIGR4) GN = ply PE = 3 SV = 1 − [TACY_STRPN]Q97N55 75 27 392 729.12 Secreted 45 kd protein Secretory OS =Streptococcus pneumoniae (released) serotype 4 (strain ATCC BAA-334/(with CS) TIGR4) GN = usp45 PE = 4 SV = 1 − [Q97N55_STRPN] Q97RY6 42.173 2140 715.85 Serine protease, subtilase family LP × TG OS =Streptococcus pneumoniae Cell-wall serotype 4 (strain ATCC BAA-334/anchored TIGR4) GN = SP_0641 PE = 4 SV = 1 − [Q97RY6_STRPN] P64030 68.8432 398 684.61 Elongation factor Tu OS = Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = tuf PE = 3 SV = 1− [EFTU_STRPN] Q97NQ7 66.42 61 1203 683.01 DNA-directed RNA polymerasesubunit Intracellular beta OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = rpoB PE = 3 SV = 1 − [RPOB_STRPN]Q97PE6 77.38 35 442 645.41 Sugar ABC transporter, sugar-binding Lipidprotein OS = Streptococcus pneumoniae anchored serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_1683 PE = 4 SV = 1 − [Q97PE6_STRPN] Q97PT1 74.5945 551 641.52 Uncharacterized protein Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1518 PE = 4SV = 1 − [Q97PT1_STRPN] Q97RQ0 81.35 26 386 599.79 Branched-chain aminoacid ABC Lipid transporter, amino acid-binding anchored protein OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =livJ PE = 4 SV = 1 − [Q97RQ0_STRPN] Q97SV2 62.45 19 277 575.15 50 Sribosomal protein L2 OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = rplB PE = 3 SV = 1 −[RL2_STRPN] Q97R51 74.12 32 313 574.61 Foldase protein PrsA OS = LipidStreptococcus pneumoniae serotype 4 anchored (strain ATCC BAA-334/TIGR4)GN = prsA PE = 3 SV = 1 − [PRSA_STRPN] Q97SV5 59.62 17 208 571.92 50 Sribosomal protein L3 OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = rpIC PE = 3 SV = 1 −[RL3_STRPN] Q04707 64.12 38 719 564.47 Penicillin-binding protein 1 A OS= N- Streptococcus pneumoniae serotype 4 terminally (strain ATCCBAA-334/TIGR4) anchored GN = ponA PE = 1 SV = 2 − [PBPA_STRPN] (No CS)Q97NL1 88.36 26 335 546.25 Glyceraldehyde-3-phosphate Intracellulardehydrogenase OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = gap PE = 3 SV = 1 − [Q97NL1_STRPN] Q97PD6 64.87 46837 501.57 Protein translocase subunit SecA 1 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =secA1 PE = 3 SV = 1 − [SECA1_STRPN] P35592 75.61 44 660 500.22Oligopeptide-binding protein AliA OS = Lipid Streptococcus pneumoniaeserotype 4 anchored (strain ATCC BAA-334/TIGR4) GN = aliA PE = 3 SV = 4− [ALIA_STRPN] P0A4G2 72.49 21 309 482.72 Manganese ABC transporterLipid substrate-binding lipoprotein OS = anchored Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = psaA PE = 1 SV =1 − [MITSA_STRPN] Q54970 61.59 38 591 482.11 Pyruvate oxidase OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = spxB PE = 3 SV = 2 − [POXB_STRPN] Q97R09 68.62 21341 442.02 Iron-compound ABC transporter, iron Lipid compound-bindingprotein OS = anchored Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1032 PE = 1 SV = 1 − [Q97R09_STRPN] Q97PU3 67.6331 278 434.28 Amino acid ABC transporter, amino Lipid acid-bindingprotein OS = Streptococcus anchored pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = aatB PE = 4 SV = 1 − [Q97PU3_STRPN] P0A4C3 72.35 20217 427.97 30 S ribosomal protein S3 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpsC PE = 3 SV =1 − [RS3_STRPN] Q97T39 36.42 28 744 407.46 Pneumococcal surface proteinA N- OS = Streptococcus pneumoniae terminally serotype 4 (strain ATCCBAA-334/ anchored TIGR4) GN = pspA PE = 4 SV = 1 − (with CS)[Q97T39_STRPN] I6L8V7 41.79 28 627 381.01 Choline binding protein E OS =Secretory Streptococcus pneumoniae serotype 4 (released) (strain ATCCBAA-334/TIGR4) (with CS) GN = cbpE PE = 4 SV = 1 − [16L8V7_STRPN] P0A47554.74 11 137 354.71 50 S ribosomal protein L16 OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =rpIP PE = 1 SV = 1 − [RL16_STRPN] P66112 51.26 9 119 346.49 50 Sribosomal protein L20 OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = rpIT PE = 3 SV = 1 −[RL20_STRPN] Q97SV4 56.04 14 207 345.01 50 S ribosomal protein L4 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpID PE = 3 SV = 1 − [RL4_STRPN] Q97T12 78.99 26 276332.96 ABC transporter, substrate-binding Lipid protein OS =Streptococcus pneumoniae anchored serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0148 PE = 4 SV = 1 − [Q97T12_STRPN] Q97N74 47.47 31 693320.12 Choline binding protein A OS = N- Streptococcus pneumoniaeserotype 4 terminally (strain ATCC BAA-334/TIGR4) anchored GN = cbpA PE= 1 SV = 1 − (with CS) [Q97N74_STRPN] Q97QS2 58.06 22 434 315.73 EnolaseOS = Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = eno PE = 1 SV = 1 − [ENO_STRPN] Q97S57 49.58 34 958315.37 Translation initiation factor IF-2 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =infB PE = 3 SV = 1 − [IF2_STRPN] Q97T11 75 27 284 313.6 Lipoprotein OS =Streptococcus Lipid pneumoniae serotype 4 (strain ATCC anchoredBAA-334/TIGR4) GN = SP_0149 PE = 3 SV = 1 − [Q97T11_STRPN] P66359 55.1210 127 299.97 30 S ribosomal protein S11 OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =rpsK PE = 3 SV = 1 − [RS11_STRPN] Q97PT6 68.8 24 468 285.18 ATP synthasesubunit beta OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = atpD PE = 3 SV = 1 − [ATPB_STRPN]Q97SP2 60.24 18 332 279.63 PTS system, mannose-specific IIABIntracellular components OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = manL PE = 4 SV = 1 − [Q97SP2_STRPN]Q97RZ7 60.08 15 238 278.06 Uncharacterized protein OS = LipidStreptococcus pneumoniae serotype 4 anchored (strain ATCC BAA-334/TIGR4)GN = SP_0629 PE = 1 SV = 1 − [Q97RZ7_STRPN] Q97NL3 55.13 34 731 275.05Penicillin-binding protein 2A OS = N- Streptococcus pneumoniae serotype4 terminally (strain ATCC BAA-334/TIGR4) anchored GN = pbp2A PE = 4 SV =1 − (No CS) [Q97NL3_STRPN] Q97PE1 62.92 29 445 269.34 ABC transporter,substrate-binding Lipid protein OS = Streptococcus anchored pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1690 PE = 4 SV = 1 −[Q97PE1_STRPN] Q97SX2 81.48 19 189 268.65 Uncharacterized protein OS =Lipid Streptococcus pneumoniae serotype 4 anchored (strain ATCCBAA-334/TIGR4) GN = SP_0191 PE = 1 SV = 1 − [Q97SX2_STRPN] Q97RN2 67.7915 267 248.33 Peptidyl-prolyl cis-trans isomerase, Lipidcyclophilin-type OS = Streptococcus anchored pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = SP_0771 PE = 3 SV = 1 − [Q97RN2_STRPN]P65887 62.62 25 428 243.93 Adenylosuccinate synthetase OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = purA PE = 1 SV = 1 − [PURA_STRPN] Q97QH2 33.7 23 721242.51 Amino acid ABC transporter, amino Multi- acid-bindingprotein/permease trans- protein OS = Streptococcus membrane pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1241 PE = 1 SV = 1 −[Q97QH2_STRPN] O69076 50.46 25 652 240.51 ATP-dependent zincmetalloprotease Multi- FtsH OS = Streptococcus pneumoniae trans-serotype 4 (strain ATCC BAA-334/ membrane TIGR4) GN = ftsH PE = 3 SV = 3− [FTSH_STRPN] Q97N37 62.6 22 393 236.55 Serine protease OS = -Streptococcus N- pneumoniae serotype 4 (strain ATCC terminallyBAA-334/TIGR4) GN = SP_2239 PE = 1 anchored SV = 1 − [Q97N37_STRPN] (NoCS) Q97T63 60.29 26 491 232.33 ABC transporter, substrate-binding Lipidprotein OS = - Streptococcus anchored pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0092 PE = 4 SV = 1 − [Q97T63_STRPN] Q97NQ0 68.7919 330 230.07 Aspartate--ammonia ligase OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = asnA PE = 3 SV =1 − [ASNA_STRPN] Q97SV1 86.67 12 180 229.35 50 S ribosomal protein L5 OS= Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rplE PE = 3 SV = 1 − [RL5_STRPN] Q97Q37 69.37 18 271227.84 Amino acid ABC transporter, amino Lipid acid-binding protein OS =anchored Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4)GN = SP_1394 PE = 4 SV = 1 − [Q97Q37_STRPN] P66907 42.63 9 380 223.83Queuine tRNA-ribosyltransferase OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = tgt PE = 3 SV = 1− [TGT_STRPN] P59205 43.77 20 658 217.64 Putative endo-beta-N- Secretoryacetylglucosaminidase OS = (released) Streptococcus pneumoniae (strain(with CS) ATCC BAA-334/TIGR4) GN = lytB PE = 1 SV = 1 − [LYTB_STRPN]Q97SC6 49.35 27 774 216.35 Formate acetyltransferase OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = pflPE = 4 SV = 1 − [Q97SC6_STRPN] Q97NB5 42.19 22 621 215.44 Cholinebinding protein PcpA OS = Secretory Streptococcus pneumoniae serotype 4(released) (strain ATCC BAA-334/TIGR4) (with CS) GN = pcpA PE = 1 SV = 1− [Q97NB5_STRPN] P0A2U8 49.58 19 355 215.01 Oligopeptide transportATP-binding Intracellular protein AmiE OS = -- Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = amiE PE = 3 SV = 1 −[AMIE_STRPN] Q97PA9 39 21 659 212.01 Serine/threonine-protein kinaseStkP Intracellular/ OS = - Streptococcus pneumoniae TMH serotype 4(strain ATCC BAA-334 start TIGR4) GN = stkP PE = 1 SV = 1 − AFTER 60[STKP2_STRPN] Q97RF9 56.25 22 400 204.01 Ribosomal protein S1 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpsA PE = 4 SV = 1 − [Q97RF9_STRPN] Q97SP4 43.89 12303 202.88 PTS system, mannose-specific IID Multi- component OS = -Streptococcus trans- pneumoniae serotype 4 (strain ATCC membraneBAA-33/TIGR4) GN = SP_0282 PE = 4 SV = 1 − [Q97SP4_STRPN] P0A4A7 58.3911 137 202.86 30 S ribosomal protein S12 OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =rpsL PE = 3 SV = 1 − [RS12_STRPN] Q97Q31 55.82 13 292 202.43Phosphate-binding protein PstS 1 OS = Lipid Streptococcus pneumoniaeserotype 4 anchored (strain ATCC BAA-334/TIGR4) GN = pstS1 PE = 1 SV = 1− [PSTS1_STRPN] Q97N56 69.11 20 259 201.44 30 S ribosomal protein S2 OS= Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpsB PE = 3 SV = 1 − [RS2_STRPN] Q97RC6 48.66 14 335201.21 ATP-dependent 6- Intracellular phosphofructokinase OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =pfkA PE = 3 SV = 1 − [PFKA_STRPN] Q97QX5 68.02 15 344 200.45Uncharacterized protein OS = Secretory Streptococcus pneumoniae serotype4 (released) (strain ATCC BAA-334/TIGR4) (with CS) GN = SP_1069 PE = 1SV = 1 − [Q97QX5_STRPN] Q97NM6 60.64 17 404 197.5 Aminotransferase,class I OS = Intracellular Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = SP_1994 PE = 4 SV = 1 − [Q97NM6_STRPN] Q97N6965.37 19 335 194.36 Putative ABC transporter, substrate- Lipid bindingprotein OS = -Streptococcus anchored pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_2197 PE = 4 SV = 1 − [Q97N69_STRPN] Q97NS5 60.9516 338 193.02 Putative transcriptional regulator OS = SecretoryStreptococcus pneumoniae serotype 4 (released) (strain ATCCBAA-334/TIGR4) (with CS) GN = SP_1942 PE = 4 SV = 1 − [Q97NS5_STRPN]P0A4M7 38.35 20 498 189.98 Oligopeptide transport system Multi- permeaseprotein AmiC OS = = trans- Streptococcus pneumoniae serotype 4 membrane(strain ATCC BAA-334/TIGR4) GN = amiC PE = 3 SV = 1 − [AMIC_STRPN]Q97QD2 56.41 20 523 188.15 Signal recognition particle protein OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = ffh PE = 1 SV = 1 − [Q97QD2_STRPN] P14677 49.87 26750 185.33 Penicillin-binding protein 2 × OS = = N- Streptococcuspneumoniae serotype 4 terminally (strain ATCC BAA-334/TIGR4) anchored GN= pbpX PE = 1 SV = 2 − [PBPX_STRPN] (No CS) Q97NQ3 51.88 14 345 183.26Uncharacterized protein OS = N- Streptococcus pneumoniae serotype 4terminally (strain ATCC BAA-334/TIGR4) anchored GN = SP_1967 PE = 4 SV =1 − (No CS) [Q97NQ3_STRPN] Q97RW9 59.34 16 332 182.68 Putativepneumococcal surface Lipid protein OS = - Streptococcus anchoredpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0667 PE = 1 SV= 1 − [Q97RW9_STRPN] Q97RL9 66.17 16 399 180.86 Uncharacterized proteinOS = N - Streptococcus pneumoniae serotype 4 terminally (strain ATCCBAA-334/TIGR4) anchored GN = SP_0785 PE = 4 SV = 1 − (No CS)[Q97RL9_STRPN] P0A3M9 47.26 14 328 180.34 L-lactate dehydrogenase OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = Idh PE = 3 SV = 2 − [LDH_STRPN] Q97PT4 39.72 17 501180.17 ATP synthase subunit alpha OS = Multi- Streptococcus pneumoniaeserotype 4 trans- (strain ATCC BAA-334/TIGR4) membrane GN = atpA PE = 3SV = 1 − [ATPA_STRPN] Q97QH1 67.48 9 246 175.25 Amino acid ABCtransporter, ATP- Intracellular binding protein OS = - Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1242 PE = 3 SV= 1 − [Q97QH1_STRPN] Q97NX9 72.9 19 321 174.95 Iron-compound ABCtransporter, iron- Lipid compound-binding protein OS = anchoredStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_1872 PE = 4 SV = 1 − [Q97NX9_STRPN] Q97QC6 66.09 14 115 173.94 50 Sribosomal protein L19 OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = rplS PE = 3 SV = 1 −[RL19_STRPN] Q97N99 63.14 18 274 172.57 SPFH domain/Band 7 family OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_2156 PE = 4 SV = 1 − [Q97N99_STRPN] Q97PW1 50.9717 463 169.74 Peptidoglycan N-acetylglucosamine N- deacetylase A OS =-Streptococcus terminally pneumoniae serotype 4 (strain ATCC anchoredBAA-334/TIGR4) GN = pgdA PE = 4 (No CS) SV = 1 − [Q97PW1_STRPN] Q97SE136.4 14 555 169.45 Uncharacterized protein OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_0443 PE = 1 SV = 1 − [Q97SE1_STRPN] Q97PM5 59.04 16 376 168.92 SugarABC transporter, ATP-binding Intracellular protein OS = -Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = msmK PE = 4 SV =1 − [Q97PM5_STRPN] Q97SN5 85.81 15 148 165.86 50 S ribosomal protein L13OS = Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rplM PE = 3 SV = 1 − [Q97SN5_STRPN] Q97PQ2 28.88 18914 162.85 Cation-transporting ATPase, E1-E2 Multi- family OS =-Streptococcus pneumoniae trans- serotype 4 (strain ATCC BAA-334/membrane TIGR4) GN = SP_1551 PE = 3 SV = 1 − [Q97PQ2_STRPN] Q97NE4 32.1621 821 161.61 Penicillin-binding protein 1 B OS = N- Streptococcuspneumoniae serotype 4 terminally (strain ATCC BAA-334/TIGR4) anchored GN= pbp1B PE = 4 SV = 1 − (No CS) [Q97NE4_STRPN] Q97Q62 54.48 20 424161.35 Psr protein OS = -Streptococcus Intracellular/ pneumoniaeserotype 4 (strain ATCC TMH BAA-334/TIGR4) GN = SP_1368 PE = 4 start SV= 1 − [Q97Q62_STRPN] AFTER 60 P18766 50 13 308 160.11 Oligopeptidetransport ATP-binding Intracellular protein AmiF OS = -Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = amiF PE = 3 SV =2 − [AMIF_STRPN] I6L8U0 42.39 16 394 155.34 UDP-N-acetylglucosamine 2-Intracellular epimerase OS = - Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = cps4L PE = 3 SV = 1 − [I6L8U0_STRPN]P0CC08 55.21 15 288 154.74 Acetyl-coenzyme A carboxylase Intracellularcarboxyl transferase subunit beta OS = Streptococcus pneumoniae serotype4 (strain ATCC BAA-334/TIGR4) GN = accD PE = 1 SV = 1 − [ACCD_STRPN]Q97RG9 51.27 21 511 154.04 Sugar ABC transporter, ATP-bindingIntracellular protein OS = -Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = SP_0846 PE = 3 SV = 1 − [Q97RG9_STRPN] Q97SJ450.28 17 360 153.5 Uncharacterized protein OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_0355 PE = 4 SV = 1 − [Q97SJ4_STRPN] P72524 33.82 22 822 148.92 DNAgyrase subunit A OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = gyrA PE = 3 SV = 3 − [GYRA_STRPN]P66565 56.16 17 203 148.91 30 S ribosomal protein S4 OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =rpsD PE = 3 SV = 1 − [RS4_STRPN] P0A4G0 41.1 20 652 146.87Oligopeptide-binding protein AliB Lipid OS = = -Streptococcus pneumoniaeanchored serotype 4 (strain ATCC BAA-334/ TIGR4) GN = aliB PE = 3 SV = 1− [ALIB_STRPN] Q97SG0 57.78 18 424 144.79 Serine--tRNA ligase OS =-Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = serS PE = 3 SV = 1 − [SYS_STRPN] Q97PF9 51.79 20 419144.46 Cell division protein FtsZ OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = ftsZ PE = 3 SV =1 − [Q97PF9_STRPN] Q97Q34 64.04 17 267 144.16 Phosphate importATP-binding protein Intracellular PstB 2 OS = - Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = pstB2 PE = 3 SV = 1 −[PSTB2_STRPN] I6L8N1 47.25 14 455 141.81 Acetyl-CoA carboxylase, biotinIntracellular carboxylase OS = -Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = accC PE = 4 SV = 1 − [I6L8N1_STRPN]Q97SR1 45.54 19 617 140.84 Proline--tRNA ligase OS = = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =proS PE = 3 SV = 1 − [SYP_STRPN] Q97SI4 40.88 11 340 140.1 Cholinebinding protein C OS = N- Streptococcus pneumoniae serotype 4 terminally(strain ATCC BAA-334/TIGR4) anchored GN = cbpC PE = 4 SV = 1 −[Q97SI4_STRPN] (with CS) P0A4D7 40.84 17 524 138.63 DEAD-boxATP-dependent RNA Intracellular helicase CshA OS = - Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = cshA PE = 3 SV =1 − [CSHA_STRPN] Q97RN3 44.44 17 513 136.66 ABC transporter, ATP-bindingprotein Intracellular OS = -Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = SP_0770 PE = 1 SV = 1 − [Q97RN3_STRPN] Q97PR043.62 17 447 136.28 Asparagine--tRNA ligase OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =asnS PE = 1 SV = 1 − [SYN_STRPN] P22976 40.73 18 658 132.04 Probabletransketolase OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = tkt PE = 3 SV = 2 − [TKT_STRPN] Q97PX134.2 16 459 132 NADH oxidase OS = = -Streptococcus Intracellularpneumoniae serotype 4 (strain ATCO BAA-334/TIGR4) GN = nox PE = 4 SV = 1− [Q97PX1_STRPN] O05703 50.5 17 501 129.61 Zinc-binding lipoprotein AdcAOS = Lipid Streptococcus pneumoniae serotype 4 anchored (strain ATCCBAA-334/TIGR4) GN = adcA PE = 3 SV = 4 − [ADCA_STRPN] P66095 52.4 12 229128.44 50 S ribosomal protein L1 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rplA PE = 3 SV =1 − [RL1_STRPN] Q97RP4 55.65 12 230 127.75 Cell division ABCtransporter, ATP- Intracellular binding protein FtsE OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = ftsE PE = 3 SV =1 − [Q97RP4_STRPN] Q97RP3 37.66 12 308 126.99 Cell division protein FtsXOS = Multi- Streptococcus pneumoniae serotype 4 trans- (strain ATCCBAA-334/TIGR4) membrane GN = ftsX PE = 3 SV = 1 − [Q97RP3_STRPN] Q97N7238.4 23 810 122.4 ATP-dependent Clp protease, ATP- Intracellular bindingsubunit OS = -Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_2194 PE = 3 SV = 1 − [Q97N72_STRPN] Q97Q67 43.914 344 121.11 Conserved domain protein OS = Multi- Streptococcuspneumoniae serotype 4 trans- (strain ATCC BAA-334/TIGR4) membrane GN =SP_1363 PE = 4 SV = 1 − [Q97Q67_STRPN] P95830 50.79 15 378 120.38Chaperone protein DnaJ OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = dnaJ PE = 1 SV = 2 −[DNAJ_STRPN] Q97SJ6 53.04 8 230 119.51 Capsular polysaccharidebiosynthesis Multi- protein CpsC OS = -Streptococcus trans- pneumoniaeserotype 4 (strain ATCC membrane BAA-334/TIGR4) GN = cpsC PE = 3 SV = 1− [CPSC_STRPN] P63373 71.03 13 252 116.79 Phosphate import ATP-bindingprotein Intracellular PstB 1 OS = -Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = pstB1 PE = 3 SV = 1 − [PSTB1_STRPN]I6L8Q3 32.35 12 340 114.44 Choline binding protein F OS = N-Streptococcus pneumoniae serotype 4 terminally (strain ATCCBAA-334/TIGR4) anchored GN = cbpF PE = 4 SV = 1 − [I6L8Q3_STRPN] (withCS) Q97RS8 62.96 16 378 110.09 Lactate oxidase OS = -StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =IctO-2 PE = 4 SV = 1 − [Q97RS8_STRPN] Q97N53 70.96 11 272 109.3 Cellshape-determining protein MreC N- OS = -Streptococcus pneumoniaeterminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = mreC PE= 3 SV = 1 − (No CS) [Q97N53_STRPN] P0A3M5 41.32 18 680 109.25Penicillin-binding protein 2 B OS = N- Streptococcus pneumoniae serotype4 terminally (strain ATCC BAA-334/TIGR4) anchored GN = penA PE = 3 SV =1 − [PBP2_STRPN] (No CS) P0A4S1 49.15 9 293 109 Fructose-bisphosphatealdolase OS = Intracellular Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = fba PE = 3 SV = 1 − [ALF_STRPN] Q97RK0 29.57 13575 107.28 Septation ring formation regulator N- EzrA OS = -Streptococcus pneumoniae terminally serotype 4 (strain ATCC BAA-334/anchored TIGR4) GN = ezrA PE = 3 SV = 1 − (No CS) [EZRA_STRPN] Q97SI960.55 7 109 107.21 Cell cycle protein GpsB OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =gpsB PE = 3 SV = 1 − [GPSB_STRPN] P63413 36.36 12 396 106.75 Acetatekinase OS = - Streptococcus Intracellular pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = ackA PE = 3 SV = 1 − [ACKA_STRPN] P0A2Z2 59.7610 164 105.18 ATP synthase subunit b OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = atpF PE = 3 SV =1 − [ATPF_STRPN] Q97SQ4 39.74 8 156 103.23 30 S ribosomal protein S7 OS= Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpsG PE = 3 SV = 1 − [RS7_STRPN] Q97QK5 48.6 19 607101.3 Elongation factor 4 OS = - Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = lepA PE = 3 SV = 1 −[LEPA_STRPN] P65239 39.75 11 322 100.54 Ribose-phosphatepyrophosphokinase Intracellular 1 OS = -Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = prs1 PE = 3 SV = 1 −[KPRS1_STRPN] P95829 36.41 14 607 100.51 Chaperone protein Dnak OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = dnak PE = 3 SV = 2 − [DNAK_STRPN] I6L8U5 55.47 11411 100.01 3-oxoacyl-[acyl-carrier-protein] Intracellular synthase 2 OS= - Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =fabF PE = 3 SV = 1 − [16L8U5_STRPN] Q97PV1 21.8 6 289 93.98 Putativeglycerol uptake facilitator Multi- protein OS = -Streptococcus trans-pneumoniae serotype 4 (strain ATCC membrane BAA-334/TIGR4) GN = SP_1491PE = 3 SV = 1 − [Q97PV1_STRPN] Q2MGH6 15.73 18 1767 93.68Endo-alpha-N-acetylgalactosaminidase LP × TG OS = -Streptococcuspneumoniae Cell-wall anchored serotype 4 (strain ATCC BAA-334/ TIGR4) GN= SP_0368 PE = 1 SV = 1 − [GH101_STRPN] Q97SQ9 36.05 13 602 92.56Glutamine--fructose-6-phosphate Intracellular aminotransferase[isomerizing] OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = glmS PE = 3 SV = 3 − [GLMS_STRPN] Q97P40 53.96 9 20292.31 Putative general stress protein 24 OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_1804 PE = 4 SV = 1 − [Q97P40_STRPN] Q97QE4 25.33 18 1058 90.95Carbamoyl-phosphate synthase large Intracellular chain OS = -Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =carB PE = 3 SV = 1 − [CARB_STRPN] I6L8Q9 40.04 11 457 90.76 Celldivision protein ftsA OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = ftsA PE = 3 SV = 1 −[I6L8Q9_STRPN] I6L8S9 45.52 10 413 90.26 D-alanyl-D-alaninecarboxypeptidase N- OS = -Streptococcus pneumoniae terminally serotype 4(strain ATCC BAA-334/ anchored TIGR4) GN = dacA PE = 3 SV = 1 − (withCS) [I6L859_STRPN] Q97Q48 27.52 18 872 89.81 Alanine--tRNA ligase OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = alaS PE = 3 SV = 1 − [SYA_STRPN] Q97QD5 64.52 12 18689.81 LemA protein OS = -Streptococcus N- pneumoniae serotype 4 (strainATCC terminally BAA-334/TIGR4) GN = lemA PE = 1 anchored SV = 1 −[Q97QD5_STRPN] (No CS) Q97NW2 48.69 13 419 88.69 Sugar ABC transporter,sugar-binding Lipid protein OS = -Streptococcus anchored pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = rafE PE = 1 SV = 1 −[Q97NW2_STRPN] I6L8N0 43.01 14 365 88.6 UDP-N-acetylglucosamine-2-Intracellular epimerase OS = - Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = cps4l PE = 3 SV = 1 − [I6L8N0_STRPN]Q97QW4 36.47 13 425 87.93 Uncharacterized protein OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_1083 PE = 4 SV = 1 − [Q97QW4_STRPN] Q9FBB7 58.43 12 255 86.89Acetyl-coenzyme A carboxylase Intracellular carboxyl transferase subunitalpha OS = -Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = accA PE = 1 SV = 1 − [ACCA_STRPN] P67266 42.42 4 99 86.55Nucleoid-associated protein SP _1102 Intracellular OS = -Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1102 PE = 1SV = 1 − [Y1102_STRPN] Q97QM4 24.06 13 719 86.48Ribonucleoside-diphosphate Intracellular reductase OS = -Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = nrdE PE = 3 SV =1 − [Q97QM4_STRPN] Q97PF5 44.17 8 283 85.34 Putativephosphosugar-binding Intracellular transcriptional regulator OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_1674 PE = 4 SV = 1 − [Q97PF5_STRPN] P0A3Y3 50.87 10 230 84.952,3-bisphosphoglycerate-dependent Intracellular phosphoglycerate mutaseOS = Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN= gpmA PE = 3 SV = 1 − [GPMA_STRPN] Q97NE1 40.78 4 103 84.35Uncharacterized protein OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_2102 PE = 4 SV = 1 −[Q97NE1_STRPN] P67282 35.58 16 534 83.04 Ribonuclease Y OS =-Streptococcus N- pneumoniae serotype 4 (strain ATCC terminallyBAA-334/TIGR4) GN = rny PE = 3 SV = 1 − anchored [RNY_STRPN] (No CS)Q97QX6 22.61 14 898 82.38 Phosphoenolpyruvate carboxylase IntracellularOS = -Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4)GN = ppc PE = 3 SV = 1 − [CAPP_STRPN] Q97SD1 32.63 12 521 81.72 Aminoacid ABC transporter, amino Multi- acid-binding protein/permease trans-protein OS = -Streptococcus membrane pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0453 PE = 4 SV = 1 − [Q97SD1_STRPN] P66419 50.568 89 78.12 30 S ribosomal protein S14 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rpsN PE = 3 SV =1 − [RS14_STRPN] P61182 61.4 8 114 77.58 50 S ribosomal protein L22 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rplV PE = 1 SV = 1 − [RL22_STRPN] P66278 33.33 4 6677.34 50 S ribosomal protein L35 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rpml PE = 3 SV =1 − [RL35_STRPN] Q97R14 34.57 10 324 76.27 Uncharacterized protein OS =Secretory Streptococcus pneumoniae serotype 4 (released) (strain ATCCBAA-334/TIGR4) (with CS) GN = SP_1027 PE = 4 SV = 1 − [Q97R14_STRPN]Q97T52 22.4 12 616 70.86 Putative capsular polysaccharide Multi-biosynthesis protein OS = trans- Streptococcus pneumoniae serotype 4membrane (strain ATCC BAA-334/TIGR4) GN = SP_0103 PE = 4 SV = 1 −[Q97T52_STRPN] Q97SW2 26.53 13 735 70.68 Anaerobic ribonucleoside-Intracellular triphosphate reductase OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = nrdD PE = 4 SV = 1 −[Q97SW2_STRPN] Q97SE4 32.3 12 514 70.62 Peptide chain release factor 3OS = Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = prfC PE = 3 SV = 1 − [RF3_STRPN] Q97PC4 27.24 9 49268.5 ABC transporter, ATP-binding protein Multi- OS = - Streptococcuspneumoniae trans- serotype 4 (strain ATCC BAA-334/ membrane TIGR4) GN =SP_1715 PE = 4 SV = 1 − [Q97PC4_STRPN] I6L8V8 47.33 7 243 68.483-oxoacyl-[acyl-carrier protein] Intracellular reductase OS = -Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =fabG PE = 3 SV = 1 − [I6L8V8_STRPN] Q97S28 21.17 11 737 68.26Polyribonucleotide Intracellular nucleotidyltransferase OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = pnpPE = 3 SV = 1 − [PNP_STRPN] Q97RC4 41.38 9 290 67.69 Uncharacterizedprotein OS = Lipid Streptococcus pneumoniae serotype 4 anchored (strainATCC BAA-334/TIGR4) GN = SP_0899 PE = 4 SV = 1 − [Q97RC4_STRPN] Q97S8621.65 7 448 65.38 Glutamine synthetase, type I OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =glnA PE = 3 SV = 1 − [Q97S86_STRPN] Q97NH4 41.48 9 352 65.13 Alcoholdehydrogenase, zinc- Intracellular containing OS = - Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_2055 PE = 3 SV= 1 − [Q97NH4_STRPN] P0A495 65.79 3 38 64.65 50 S ribosomal protein L36OS = Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpmJ PE = 3 SV = 1 − [RL36_STRPN] Q97ND6 25.72 13587 64.48 Aspartate--tRNA ligase OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = aspS PE = 3 SV =1 − [SYD_STRPN] P72525 18.1 10 823 63.42 DNA topoisomerase 4 subunit AOS = Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = parC PE = 1 SV = 3 − [PARC_STRPN] Q97PH2 23.11 12740 63.08 GTP pyrophosphokinase OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = relA PE = 3 SV =1 − [Q97PH2_STRPN] Q97T46 32.63 4 95 63.06 Putative bacteriocin OS =Secretory Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0109 PE = 4 SV = 1 − [Q97T46_STRPN] I6L8V0 42.1711 351 60.21 Capsular polysaccharide biosynthesis Intracellular proteinCps4J OS = - Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = cap4J PE = 4 SV = 1 − [I6L8V0_STRPN] Q97RQ6 23.84 6281 60.06 DegV domain-containing protein Intracellular SP_0742 OS = -Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_0742 PE = 3 SV = 1 − [Y742_STRPN] Q97SR2 35.08 9 419 60.03 Putativezinc metalloprotease Multi- SP_0263 OS = - Streptococcus trans-pneumoniae serotype 4 (strain ATCC membrane BAA-334/TIGR4) GN = SP_0263PE = 3 SV = 1 − [Y263_STRPN] Q97PB8 15.63 8 627 59.73 PTS system IIABCcomponents OS = Multi- Streptococcus pneumoniae serotype 4 trans-(strain ATCC BAA-334/TIGR4) membrane GN = SP_1722 PE = 4 SV = 1 −[Q97PB8_STRPN] Q97QE6 41.47 9 340 58.95 Alcohol dehydrogenase, zinc-Intracellular containing OS = -Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = SP_1270 PE = 4 SV = 1 − [Q97QE6_STRPN]Q97P07 34.56 10 408 58.93 Putative capsular polysaccharide Intracellularbiosynthesis protein OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = SP_1837 PE = 3 SV = 1 − [Q97P07_STRPN] Q97T0940.23 9 353 57.99 Methionine import ATP-binding Intracellular proteinMetN OS = -Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = metN PE = 3 SV = 1 − [METN_STRPN] Q97NP5 18.83 7 30857.43 Membrane protein insertase YidC 1 Multi- OS = -Streptococcuspneumoniae trans- serotype 4 (strain ATCC BAA-334/ membrane TIGR4) GN =yidC1 PE = 3 SV = 1 − [YIDC1_STRPN] P0A451 41.49 12 388 57.14 ProteinRecA OS = -Streptococcus Intracellular pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = recA PE = 2 SV = 1 − [RECA_STRPN] Q97SU7 51.125 178 56.93 50 S ribosomal protein L6 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rplF PE = 3 SV =1 − [RL6_STRPN] Q97NE2 18.49 9 687 56.54 Cation-transporting ATPase,E1-E2 Intracellular family OS = -Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = SP_2101 PE = 3 SV = 1 − [Q97NE2_STRPN]Q97SN4 32.31 6 130 56.22 30 S ribosomal protein S9 OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =rpsl PE = 3 SV = 1 − [RS9_STRPN] P66392 51.24 7 121 56.15 30 S ribosomalprotein S13 OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = rpsM PE = 3 SV = 1 − [RS13_STRPN]Q97S10 23.51 11 553 55.98 Ribonuclease J OS = - StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rnjPE = 3 SV = 1 − [Q97S10_STRPN] Q97PG0 20.99 5 262 55.94 Cell divisionprotein DivIVA OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = divlVA PE = 4 SV = 1 − [Q97PG0_STRPN]P35597 17.61 9 778 55.84 Probable cation-transporting ATPase Multi- exp7OS = - Streptococcus pneumoniae trans- serotype 4 (strain ATCC BAA-334/membrane TIGR4) GN = exp7 PE = 3 SV = 2 − [EXP7_STRPN] Q97SJ2 43.52 12409 55.75 Capsular polysaccharide biosynthesis Intracellular proteinCps4K OS = - Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = cps4K PE = 4 SV = 1 − [Q97SJ2_STRPN] Q97PI4 29.98 12647 55.52 Threonine--tRNA ligase OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = thrS PE = 1 SV =1 − [SYT_STRPN] Q97QW8 25.81 8 434 55.41 GTPase Obg OS = - StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = obgPE = 3 SV = 1 − [OBG_STRPN] P67293 43.9 3 82 55.21 UPF0154 proteinSP_1882 OS = N- Streptococcus pneumoniae serotype 4 terminally (strainATCC BAA-334/TIGR4) anchored GN = SP_1882 PE = 3 SV = 1 − (No CS)[Y1882_STRPN] P0A4L9 20.52 10 648 54.97 DNA gyrase subunit B OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = gyrB PE = 3 SV = 1 − [GYRB_STRPN] Q97SJ8 33.2 11 49454.78 UPF0371 protein SP_0341 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0341 PE = 3 SV= 1 − [Y341_STRPN] Q54869 24.16 10 563 54.43 Arginine--tRNA ligase OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = argS PE = 1 SV = 2 − [SYR_STRPN] P35595 14.88 7 72653.64 PTS system glucose-specific EIICBA Multi- component OS =-Streptococcus trans- pneumoniae serotype 4 (strain ATCC membraneBAA-334/TIGR4) GN = exp5 PE = 3 SV = 2 − [PTG3C_STRPN] Q97SI7 22.41 7464 53.36 Mid-cell-anchored protein Z OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = mapZ PE = 1 SV =1 − [Q97SI7_STRPN] P0A4B5 55.91 7 93 52.72 30 S ribosomal protein S19 OS= Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpsS PE = 3 SV = 1 − [RS19_STRPN] Q97SU3 23.29 4 14652.09 50 S ribosomal protein L15 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rplO PE = 3 SV =1 − [RL15_STRPN] Q97RV5 27.57 11 613 51.74 Elongation factor Tu familyprotein Intracellular OS = -Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = SP_0681 PE = 4 SV = 1 − [Q97RV5_STRPN] Q97NW98.4 5 655 50.97 Trehalose PTS system, IIABC Multi- components OS =-Streptococcus trans- pneumoniae serotype 4 (strain ATCC membraneBAA-334/TIGR4) GN = SP_1884 PE = 4 SV = 1 − [Q97NW9_STRPN] Q97QE7 41.6 9262 49.97 Choline kinase OS = -Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = pck PE = 4 SV = 1 −[Q97QE7_STRPN] P67595 31.96 8 341 48.09 Tryptophan--tRNA ligase OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = trpS PE = 3 SV = 1 − [SYW_STRPN] P66581 55.49 6 16447.77 30 S ribosomal protein S5 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rpsE PE = 3 SV =1 − [RS5_STRPN] Q97R57 16.33 10 784 47.19 Ribonuclease R OS = -Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rnr PE = 3 SV = 1 − [Q97R57_STRPN] Q97RP0 21.03 7447 46.86 DEAD-box ATP-dependent RNA Intracellular helicase CshB OS = -Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =cshB PE = 3 SV = 1 − [Q97RP0_STRPN] P0A3R1 22.65 9 649 46.48 DNAmismatch repair protein HexB Intracellular OS = -Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = hexB PE = 3 SV =1 − [HEXB_STRPN] P0A3R3 18.48 11 844 45.87 DNA mismatch repair proteinHexA Intracellular OS = -Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = hexA PE = 3 SV = 1 − [HEXA_STRPN] Q97SF6 27.977 261 45.73 Enoyl-CoA hydratase/isomerase family Intracellular proteinOS = -Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN= SP_0415 PE = 4 SV = 1 − [Q97SF6_STRPN] P66708 45.98 9 311 45.6DNA-directed RNA polymerase subunit alpha Intracellular OS =-Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rpoA PE = 3 SV = 1 − [RPOA_STRPN] I6L858 28.69 9 481 45.33 Capsularpolysaccharide biosynthesis Multi- protein Cps4A OS = - Streptococcustrans- pneumoniae serotype 4 (strain ATCC membrane BAA-334/TIGR4) GN =cps4A PE = 4 SV = 1 − [I6L8S8_STRPN] Q97N43 19.51 7 492 45.08Inosine-5′-monophosphate Intracellular dehydrogenase OS = -Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = guaB PE = 3 SV =1 − [Q97N43_STRPN] I6L8W8 16.87 6 332 44.57 Choline binding protein J OS= N- Streptococcus pneumoniae serotype 4 terminally (strain ATCCBAA-334/TIGR4) anchored GN = cbpJ PE = 4 SV = 1 − [I6L8W8_STRPN] (withCS) Q2MGG2 27.62 7 467 44.4 Serine protease, subtilase family OS =Secretory Streptococcus pneumoniae serotype 4 (released) (strain ATCCBAA-334/TIGR4) (with CS) GN = SP_1954 PE = 4 SV = 1 − [Q2MGG2_STRPN]Q97SP0 42.22 7 270 44.36 Cof family protein OS = -StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_0286 PE = 4 SV = 1 − [Q97SP0_STRPN] Q97PG9 38.75 7 240 44.16Manganese ABC transporter, ATP- Intracellular binding protein OS = -Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =psaB PE = 3 SV = 1 − [Q97PG9_STRPN] Q97T72 31.67 6 221 44.14 Potassiumuptake protein, Trk family N- OS = -Streptococcus pneumoniae terminallyserotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = SP_0079 PE = 4 SV= 1 − (No CS) [Q97T72_STRPN] Q97RS9 21.98 7 496 44.07 Lysine--tRNAligase OS = - Streptococcus Intracellular pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = lysS PE = 1 SV = 2 − [SYK_STRPN] Q97PK5 27.22 2158 43.99 Uncharacterized protein OS = N- Streptococcus pneumoniaeserotype 4 terminally (strain ATCC BAA-334/TIGR4) anchored GN = SP_1604PE = 4 SV = 1 − (No CS) [Q97PK5_STRPN] Q97RV8 38.89 6 126 43.74Uncharacterized protein OS = N- Streptococcus pneumoniae serotype 4terminally (strain ATCC BAA-334/TIGR4) anchored GN = SP_0678 PE = 4 SV =1 − (No CS) [Q97RV8_STRPN] P06653 26.1 6 318 43.37 Autolysin OS =-Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = lytA PE = 1 SV = 2 − [ALYS_STRPN] P63791 27.8 8 41042.54 ATP-dependent Clp protease ATP- Intracellular binding subunit ClpXOS = Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN= clpX PE = 3 SV = 1 − [CLPX_STRPN] Q97Q36 37.33 7 217 42.45Phosphate-specific transport system Intracellular accessory protein PhoUOS = Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN= SP_1395 PE = 3 SV = 1 − [Q97Q36_STRPN] Q97R16 32.06 10 418 42.38Serine hydroxymethyltransferase OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = glyA PE = 3 SV =1 − [GLYA_STRPN] Q97NM1 38.99 9 336 42.33 Catabolite control protein AOS = Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = ccpA PE = 4 SV = 1 − [Q97NM1_STRPN] Q97PQ8 42.66 6143 40.36 Uncharacterized protein OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1545 PE = 4 SV= 1 − [Q97PQ8_STRPN] Q97SU6 59.32 7 118 40.13 50 S ribosomal protein L18OS = Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rplR PE = 3 SV = 1 − [RL18_STRPN] Q97RC9 30.77 3 15640.01 Arginine repressor OS = - Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = argR PE = 3 SV = 1 −[Q97RC9_STRPN] Q97RC5 25.15 8 501 39.74 Pyruvate kinase OS =-Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = pyk PE = 3 SV = 1 − [Q97RC5_STRPN] Q97PM1 45.04 7262 38.81 GTP-sensing transcriptional pleiotropic Intracellularrepressor CodY OS = - Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = codY PE = 1 SV = 1 − [CODY_STRPN] I6L8N6 28.91 5 21138.64 Capsular polysaccharide biosynthesis N- protein Cps4E OS = -Streptococcus terminally pneumoniae serotype 4 (strain ATCC anchoredBAA-334/TIGR4) GN = cps4E PE = 4 (No CS) SV = 1 − [I6L8N6_STRPN] Q97R3638.38 7 185 38.61 Thioredoxin family protein OS = Lipid Streptococcuspneumoniae serotype 4 anchored (strain ATCC BAA-334/TIGR4) GN = SP_1000PE = 1 SV = 1 − [Q97R36_STRPN] Q97S93 17.76 7 535 37.74 CTP synthase OS= - Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = pyrG PE = 3 SV = 1 − [PYRG_STRPN] Q97QW1 20.71 9 76337.7 DNA helicase OS = - Streptococcus Intracellular pneumoniae serotype4 (strain ATCC BAA-334/TIGR4) GN = pcrA PE = 4 SV = 1 − [Q97QW1_STRPN]P63544 51.76 6 170 37.59 Adenine phosphoribosyltransferase IntracellularOS = -Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4)GN = apt PE = 1 SV = 1 − [APT_STRPN] P65241 18.18 5 319 37.57Ribose-phosphate pyrophosphokinase Intracellular 2 OS = -Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = prs2 PE = 3 SV =1 − [KPRS2_STRPN] Q97RE5 17.08 6 650 37.17 PTS system, fructose specificIIABC Multi- components OS = -Streptococcus trans- pneumoniae serotype 4(strain ATCC membrane BAA-334/TIGR4) GN = SP_0877 PE = 4 SV = 1 −[Q97RE5_STRPN] P35596 21.38 9 608 37.12 Alpha-glycerophosphate oxidaseOS = N- Streptococcus pneumoniae serotype 4 terminally (strain ATCCBAA-334/TIGR4) anchored GN = glpO PE = 3 SV = 2 − [GLPO_STRPN] (No CS)Q97R12 19.52 7 543 37.11 Uncharacterized RNA Intracellularmethyltransferase SP_1029 OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = SP_1029 PE = 3 SV = 1 − [Y1029_STRPN]P64166 12.91 7 767 36.81 DNA translocase Ftsk OS = Multi- Streptococcuspneumoniae serotype 4 trans- (strain ATCC BAA-334/TIGR4) membrane GN =ftsK PE = 3 SV = 1 − [FTSK_STRPN] P35594 17.15 10 752 36.7 ATP-dependentClp protease ATP- Intracellular binding subunit ClpE OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = clpE PE = 3 SV =2 − [CLPE_STRPN] P64072 53.33 8 195 36.63 Probable GTP-binding proteinEngB Intracellular OS = -Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = engB PE = 3 SV = 1 − [ENGB_STRPN] Q97TC4 31.115 180 36.56 Hypoxanthine-guanine Intracellular phosphoribosyltransferaseOS = Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN= hpt PE = 3 SV = 1 − [HPRT_STRPN] Q97PU2 37.8 4 209 36.45 Amino acidABC transporter, ATP- Intracellular binding protein OS = - Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1501 PE = 3 SV= 1 − [Q97PU2_STRPN] P0A3S3 38.69 6 274 36.43 DNA-entry nuclease OS = N-Streptococcus pneumoniae serotype 4 terminally (strain ATCCBAA-334/TIGR4) anchored GN = endA PE = 1 SV = 1 − [NUCE_STRPN] (No CS)I6L8N9 25.49 8 506 36.2 Choline transporter OS = -Streptococcus Multi-pneumoniae serotype 4 (strain ATCC trans- BAA-334/TIGR4) GN = proWX PE =3 membrane SV = 1 − [16L8N9_STRPN] Q97PQ5 25.99 8 531 36.19Uncharacterized protein OS = Multi- Streptococcus pneumoniae serotype 4trans- (strain ATCC BAA-334/TIGR4) membrane GN = SP_1548 PE = 4 SV = 1 −[Q97PQ5_STRPN] Q97NI7 28.66 6 328 36.13 Putative jag protein OS =-Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_2040 PE = 4 SV = 1 − [Q97NI7_STRPN] P63742 19.327 502 36.12 Glycerol kinase OS = -Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = glpK PE = 3 SV = 1 −[GLPK_STRPN] Q97NT7 32.91 4 158 36 Uncharacterized protein OS = Multi-Streptococcus pneumoniae serotype 4 trans- (strain ATCC BAA-334/TIGR4)membrane GN = SP_1926 PE = 4 SV = 1 − [Q97NT7_STRPN] P64297 23.85 7 52035.92 GMP synthase [glutamine-hydrolyzing] Intracellular OS =-Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =guaA PE = 3 SV = 1 − [GUAA_STRPN] Q97R84 28.38 7 444 35.81Methylenetetrahydrofolate--tRNA- Intracellular(uracil-5-)-methyltransferase TrmFO OS = -Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = trmFO PE = 3 SV = 2 −[TRMFO_STRPN] Q97PK2 19.17 5 339 35.39 UDP-glucose 4-epimerase OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = galE-1 PE = 3 SV = 1 − [Q97PK2_STRPN] Q97RP6 41.53 6236 35.16 Branched-chain amino acid ABC Intracellular transporter,ATP-binding protein OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = livF PE = 3 SV = 1 − [Q97RP6_STRPN] Q97NM330.74 10 462 35.12 Cof family protein OS = -Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1997 PE = 4 SV= 1 − [Q97NM3_STRPN] Q97S34 10.86 7 801 34.77 Phenylalanine--tRNA ligasebeta Intracellular subunit OS = -Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = pheT PE = 3 SV = 1 − [SYFB_STRPN]Q97NQ4 31.38 9 427 34.05 UDP-N-acetylglucosamine 1- Intracellularcarboxyvinyltransferase 1 OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = murA1 PE = 3 SV = 1 − [MURA1_STRPN]Q97PY0 33.2 6 247 34.04 Amino acid ABC transporter, ATP- Intracellularbinding protein OS = -Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1460 PE = 3 SV = 1 − [Q97PY0_STRPN] Q97RQ3 44.5 6209 33.91 Uracil phosphoribosyltransferase OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = uppPE = 3 SV = 1 − [UPP_STRPN] Q97QP0 15.87 7 567 33.86 Dihydrolipoyldehydrogenase OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = SP_1161 PE = 4 SV = 1 − [Q97QP0_STRPN]P64062 24.77 8 436 33.49 GTPase Der OS = -Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = der PE = 3 SV = 1− [DER_STRPN] Q97N48 22.83 6 276 33.26 Uncharacterized protein OS =Intracellular/ Streptococcus pneumoniae serotype 4 TMH (strain ATCCBAA-334/TIGR4) start GN = SP_2223 PE = 4 SV = 1 − AFTER 60[Q97N48_STRPN] Q97SJ5 22.98 7 409 32.72 Capsular polysaccharidebiosynthesis Multi- protein Cps4F OS = - Streptococcus trans- pneumoniaeserotype 4 (strain ATCC membrane BAA-334/TIGR4) GN = cps4F PE = 4 SV = 1− [Q97SJ5_STRPN] P65832 36.73 6 275 32.69 Pur operon repressor OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = purR PE = 3 SV = 1 − [PURR_STRPN] P63384 12.09 9 94331.63 UvrABC system protein A OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = uvrA PE = 3 SV =1 − [UVRA_STRPN] I6L8W3 33.88 8 242 31.53 Choline transporter OS = -Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = proV PE = 3 SV = 1 − [I6L8W3_STRPN] Q9AHD2 40.97 8227 31.44 Tyrosine-protein kinase CpsD OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = cpsD PE = 3 SV =1 − [CPSD_STRPN] O08397 23.84 8 453 31.17 Chromosomal replicationinitiator Intracellular protein DnaA OS = - Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = dnaA PE = 3 SV = 2 −[DNAA_STRPN] Q97PT5 23.63 7 292 30.59 ATP synthase gamma chain OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = atpG PE = 3 SV = 1 − [ATPG_STRPN] Q97PJ1 22.09 5 24930.29 Acyltransferase family protein OS = Multi- Streptococcuspneumoniae serotype 4 trans- (strain ATCC BAA-334/TIGR4) membrane GN =SP_1624 PE = 4 SV = 1 − [Q97PJ1_STRPN] Q97NF6 14.36 6 564 30.07 ABCtransporter, ATP- Multi- binding/permease protein OS = trans-Streptococcus pneumoniae serotype 4 membrane (strain ATCC BAA-334/TIGR4)GN = SP_2075 PE = 3 SV = 1 − [Q97NF6_STRPN] Q97PW8 18.36 6 414 30.06Putative oxidoreductase OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1472 PE = 4 SV = 1 −[Q97PW8_STRPN] Q97QT0 15.61 5 474 29.86 Glyceraldehyde-3-phosphateIntracellular dehydrogenase, NADP-dependent OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = gapN PE = 3 SV =1 − [Q97QT0_STRPN] Q97PW6 12.24 6 678 29.5 Glycine--tRNA ligase betasubunit Intracellular OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = glyS PE = 3 SV = 1 − [SYGB_STRPN] Q97T93 25.938 432 29.32 Adenylosuccinate lyase OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = purB PE = 3 SV =1 − [Q97T93_STRPN] Q97RG3 5.68 2 229 29.15 Putative membrane protein OS= Multi- Streptococcus pneumoniae serotype 4 trans- (strain ATCCBAA-334/TIGR4) membrane GN = SP_0858 PE = 4 SV = 1 − [Q97RG3_STRPN]Q97TA6 12.42 3 330 28.21 Phosphate acyltransferase OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =plsX PE = 3 SV = 1 − [PLSX_STRPN] Q97TA8 19.02 6 389 27.91 Aromaticamino acid aminotransferase Intracellular OS = -Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = araT PE = 3 SV = 1 −[Q97TA8_STRPN] P0A3B7 23.99 5 346 27.48 Elongation factor Ts OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = tsf PE = 1 SV = 2 − [EFTS_STRPN] I6L8Q7 20.99 4 32427.27 Enoyl-(Acyl-carrier-protein) reductase Intracellular OS =-Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =fabK PE = 4 SV = 1 − [I6L8Q7_STRPN] Q97SG5 9.38 5 778 26.77 EndonucleaseMutS2 OS = Intracellular Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = mutS2 PE = 3 SV = 1 − [MUTS2_STRPN] Q97S79 15.45 487 26.69 Type I restriction-modification system, Intracellular Msubunit OS = -Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = hsdM PE = 4 SV = 1 − [Q97S79_STRPN] Q97SF8 12.78 4454 26.54 Aspartokinase OS = -Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0413 PE = 3 SV = 1 −[Q97SF8_STRPN] Q97S89 19.1 5 398 26.46 Phosphoglycerate kinase OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = pgk PE = 3 SV = 1 − [PGK_STRPN] Q97QH0 22.14 6 42926.44 Signal recognition particle receptor Intracellular FtsY OS =-Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =ftsY PE = 1 SV = 1 − [Q97QH0_STRPN] P0CB59 5.83 3 326 26.31Phospho-N-acetylmuramoyl- Multi- pentapeptide-transferase OS = trans-Streptococcus pneumoniae serotype 4 membrane (strain ATCC BAA-334/TIGR4)GN = mraY PE = 3 SV = 1 − [MRAY_STRPN] Q97P92 23.64 5 368 26.11GTP-binding protein OS = Intracellular Streptococcus pneumoniae serotype4 (strain ATCC BAA-334/TIGR4) GN = SP_1749 PE = 4 SV = 1 −[Q97P92_STRPN] P63733 20.06 6 359 25.98 Carbamoyl-phosphate synthasesmall Intracellular chain OS = -Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = carA PE = 3 SV = 1 − [CARA_STRPN]Q97SP1 28.61 6 339 25.89 Alcohol dehydrogenase, zinc- Intracellularcontaining OS = -Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0285 PE = 3 SV = 1 − [Q97SP1_STRPN] Q97PP2 19.924 256 25.75 Uncharacterized protein OS = Multi- Streptococcus pneumoniaeserotype 4 trans- (strain ATCC BAA-334/TIGR4) membrane GN = SP_1561 PE =1 SV = 1 − [Q97PP2_STRPN] P66339 48.04 4 102 25.74 30 S ribosomalprotein S10 OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = rpsJ PE = 3 SV = 1 − [RS10_STRPN]P0A335 13.52 6 540 25.62 60 kDa chaperonin OS = -StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =groL PE = 1 SV = 1 − [CH60_STRPN] O07344 15.69 3 204 25.56 Signalpeptidase I OS = -Streptococcus N- pneumoniae serotype 4 (strain ATCCterminally BAA-334/TIGR4) GN = lepB PE = 3 anchored SV = 2 − [LEP_STRPN](No CS) Q97PP5 41.41 3 128 25.55 Uncharacterized protein OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1558 PE = 1 SV = 1 − [Q97PP5_STRPN] Q97RP7 15.752 254 25.33 Branched-chain amino acid ABC Intracellular transporter,ATP-binding protein OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = livG PE = 3 SV = 1 − [Q97RP7_STRPN] Q97Q1213.37 5 486 24.99 Nicotinate phosphoribosyltransferase Intracellular OS= -Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =SP_1421 PE = 3 SV = 1 − [Q97Q12_STRPN] P67506 26.07 5 211 24.79 tRNA(guanine-N(7)-)- Intracellular methyltransferase OS = -Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = trmB PE = 1 SV =1 − [TRMB_STRPN] P66155 17.74 2 62 24.21 50 S ribosomal protein L28 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpmB PE = 3 SV = 1 − [RL28_STRPN] P66200 71.25 3 8023.95 50 S ribosomal protein L31 type B OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rpmE2 PE = 3 SV =1 − [RL31B_STRPN] Q97NH2 17.69 5 605 23.83 Uncharacterized protein OS =Multi- Streptococcus pneumoniae serotype 4 trans- (strain ATCCBAA-334/TIGR4) membrane GN = SP_2057 PE = 4 SV = 1 − [Q97NH2_STRPN]Q97PP4 12.89 4 450 23.67 Phosphoglucosamine mutase OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =glmM PE = 3 SV = 1 − [GLMM_STRPN] Q97R44 50.43 4 115 23.65Uncharacterized protein OS = Intracellular/ Streptococcus pneumoniaeserotype 4 TMH (strain ATCC BAA-334/TIGR4) start GN = SP_0990 PE = 4 SV= 1 − AFTER 60 [Q97R44_STRPN] Q97QB4 15.18 4 448 23.57 Glutamatedehydrogenase OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = gdhA PE = 3 SV = 1 − [Q97QB4_STRPN]Q97SW6 23.68 2 152 23.55 Uncharacterized protein OS = LipidStreptococcus pneumoniae serotype 4 anchored (strain ATCC BAA-334/TIGR4)GN = SP_0198 PE = 1 SV = 1 − [Q97SW6_STRPN] Q97PW5 16.07 4 305 23.51Glycine--tRNA ligase alpha subunit Intracellular OS = -Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = glyQ PE = 3 SV =1 − [SYGA_STRPN] Q97Q69 19.63 5 428 23.47 Homoserine dehydrogenase OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = hom PE = 3 SV = 1 − [Q97Q69_STRPN] Q97QT2 12.27 6652 23.37 DNA ligase OS = - Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = ligA PE = 3 SV = 1 −[DNLJ_STRPN] P0A4B3 33.72 3 86 23.26 30 S ribosomal protein S17 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpsQ PE = 3 SV = 1 − [RS17_STRPN] Q97NG0 14.92 4 44923.07 Glucose-6-phosphate isomerase OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = pgi PE = 3 SV = 1− [G6PI_STRPN] Q97T58 14.41 3 354 22.73 Conserved domain protein OS =Multi- Streptococcus pneumoniae serotype 4 trans- (strain ATCCBAA-334/TIGR4) membrane GN = SP_0097 PE = 4 SV = 1 − [Q97T58_STRPN]Q97P32 16.46 4 407 22.04 Tryptophan synthase beta chain OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = trpB PE = 3 SV = 1 − [TRPB_STRPN] Q97QX3 28.57 5 25221.94 ABC transporter, ATP-binding protein Intracellular OS =-Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =SP_1071 PE = 3 SV = 1 − [Q97QX3_STRPN] Q97ST6 32.03 2 128 21.84 50 Sribosomal protein L17 OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = rplQ PE = 3 SV = 1 −[RL17_STRPN] Q97RB6 8.74 5 709 21.38 Putative transcriptional regulatorOS = Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0908 PE = 4 SV = 1 − [Q97RB6_STRPN] Q97NB9 17.914 335 21.02 Uncharacterized protein OS = Multi- Streptococcus pneumoniaeserotype 4 trans- (strain ATCC BAA-334/TIGR4) membrane GN = SP_2132 PE =4 SV = 1 − [Q97NB9_STRPN] P66524 37.93 3 58 20.52 30 S ribosomal proteinS21 OS = Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpsU PE = 3 SV = 1 − [RS21_STRPN] Q97SI6 14.98 5 47420.22 6-phosphogluconate dehydrogenase, Intracellular decarboxylating OS= -Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =gnd PE = 3 SV = 1 − [Q97SI6_STRPN] Q97R47 30.45 4 266 20.09Uncharacterized protein OS = N- Streptococcus pneumoniae serotype 4terminally (strain ATCC BAA-334/TIGR4) anchored GN = SP_0987 PE = 1 SV =1 − (No CS) [Q97R47_STRPN]

TABLE 4 Proteins identified in MP from TIGR4. The accession numbersrefer to SwissProt on the date of filing. Accession # # CoveragePeptides #AAs Score Description Localization Q97RH0 76.86 46 350 2085.84Lipoprotein OS =- Streptococcus Lipid pneumoniae serotype 4 (strain ATCCanchored BAA-334/TIGR4) GN = SP_0845 PE = 4 SV = 1 - [Q97RH0_STRPN]Q97PE6 90.27 56 442 1745.9 Sugar ABC transporter, sugar-binding Lipidprotein OS =- Streptococcus anchored pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1683 PE = 4 SV = 1 - [Q97PE6_STRPN] Q97T39 61.2950 744 1569.47 Pneumococcal surface protein A OS = N- Streptococcuspneumoniae serotype 4 terminally (strain ATCC BAA-334/TIGR4) anchored GN= pspA PE = 4 SV = 1 - (with CS) [Q97T39_STRPN] P59213 82.74 62 4231500.08 Maltose/maltodextrin-binding protein Lipid OS =- Streptococcuspneumoniae anchored serotype 4 (strain ATCC BAA-334/ TIGR4) GN = malX PE= 1 SV = 1 - [MALX_STRPN] Q97N74 53.25 47 693 1294.91 Choline bindingprotein A OS = N- Streptococcus pneumoniae serotype 4 terminally (strainATCC BAA-334/TIGR4) anchored GN = cbpA PE = 1 SV = 1 - (with CS)[Q97N74_STRPN] Q97R51 87.86 45 313 1201.93 Foldase protein PrsA OS =Lipid Streptococcus pneumoniae serotype 4 anchored (strain ATCCBAA-334/TIGR4) GN = prsA PE = 3 SV = 1 - [PRSA_STRPN] Q54970 70.05 54591 1000.77 Pyruvate oxidase OS =- Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = spxB PE = 3 SV =2 - [POXB_STRPN] Q2MGF6 75.51 50 490 957.09 Lysozyme OS =- StreptococcusSecretory pneumoniae serotype 4 (strain ATCC (released) BAA-334/TIGR4)GN = lytC PE = 4 SV = 1 - (with CS) [Q2MGF6_STRPN] Q97PT1 77.13 52 551947.11 Uncharacterized protein OS = Intracellular/ Streptococcuspneumoniae serotype 4 TMH start (strain ATCC BAA-334/TIGR4) AFTER 60 GN= SP_1518 PE = 4 SV = 1 - [Q97PT1_STRPN] Q97QS2 75.35 31 434 783.92Enolase OS =- Streptococcus Intracellular pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = eno PE = 1 SV = 1 - [ENO_STRPN] P64022 76.77 48693 749.91 Elongation factor G OS =- Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = fusA PE = 3 SV =1 - [EFG_STRPN] Q04707 72.6 47 719 715.79 Penicillin-binding protein 1AOS = N- Streptococcus pneumoniae serotype 4 terminally (strain ATCCBAA-334/TIGR4) anchored GN = ponA PE = 1 SV = 2 - [PBPA_STRPN] (No CS)P0C2J9 87.47 49 471 703.79 Pneumolysin OS =- Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = ply PE = 3 SV =1 - [TACY_STRPN] Q97PE1 73.93 35 445 655.97 ABC transporter,substrate-binding Lipid protein OS =- Streptococcus anchored pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1690 PE = 4 SV = 1 -[Q97PE1_STRPN] Q97NQ8 61.8 68 1225 650.84 DNA-directed RNA polymerasesubunit Intracellular beta' OS =- Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = rpoC PE = 3 SV = 1 - [RPOC_STRPN]Q97NL1 79.1 29 335 611.32 Glyceraldehyde-3-phosphate Intracellulardehydrogenase OS =- Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = gap PE = 3 SV = 1 - [Q97NL1_STRPN] Q97T12 82.97 33276 589.52 ABC transporter, substrate-binding Lipid protein OS =-Streptococcus anchored pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4)GN = SP_0148 PE = 4 SV = 1 - [Q97T12_STRPN] Q97T63 83.3 36 491 576.45ABC transporter, substrate-binding Lipid protein OS =- Streptococcusanchored pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0092PE = 4 SV = 1 - [Q97T63_STRPN] I6L8V7 44.02 33 627 457.19 Cholinebinding protein E OS = Secretory Streptococcus pneumoniae serotype 4(released) (strain ATCC BAA-334/TIGR4) (with CS) GN = cbpE PE = 4 SV =1 - [I6L8V7_STRPN] Q97NQ7 55.61 50 1203 449.03 DNA-directed RNApolymerase subunit Intracellular beta OS =- Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpoB PE = 3 SV = 1 -[RPOB_STRPN] Q97SV2 64.62 18 277 395.51 50S ribosomal protein L2 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rplB PE = 3 SV = 1 - [RL2_STRPN] P59205 60.33 29 658388.39 Putative endo-beta-N- Secretory acetylglucosaminidase OS =(released) Streptococcus pneumoniae serotype 4 (with CS) (strain ATCCBAA-334/TIGR4) GN = lytB PE = 1 SV = 1 - [LYTB_STRPN] O69076 55.98 31652 385.64 ATP-dependent zinc metalloprotease Multi- FtsH OS =-Streptococcus pneumoniae transmembrane serotype 4 (strain ATCC BAA-334/TIGR4) GN = ftsH PE = 3 SV = 3 - [FTSH_STRPN] Q97SP2 67.47 25 332 369.57PTS system, mannose-specific IIAB Intracellular components OS =-Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =manL PE = 4 SV = 1 - [Q97SP2_STRPN] Q97T11 66.55 23 284 361.41Lipoprotein OS =- Streptococcus Lipid pneumoniae serotype 4 (strain ATCCanchored BAA-334/TIGR4) GN = SP_0149 PE = 3 SV = 1 - [Q97T11_STRPN]I6L8Q3 52.65 17 340 354.19 Choline binding protein F OS = N-Streptococcus pneumoniae serotype 4 terminally (strain ATCCBAA-334/TIGR4) anchored GN = cbpF PE = 4 SV = 1 - [I6L8Q3_STRPN] (withCS) Q97NX9 80.69 25 321 336.18 Iron-compound ABC transporter, iron-Lipid anchored compound-binding protein OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1872 PE = 4 SV = 1 -[Q97NX9_STRPN] P64030 57.29 25 398 336.06 Elongation factor Tu OS =-Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = tuf PE = 3 SV = 1 - [EFTU_STRPN] Q97NW2 64.44 25 419331.55 Sugar ABC transporter, sugar-binding Lipid protein OS =-Streptococcus anchored pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4)GN = rafE PE = 1 SV = 1 - [Q97NW2_STRPN] Q97QH2 42.86 29 721 330.61Amino acid ABC transporter, amino Multi- acid-binding protein/permeaseprotein transmembrane OS =- Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = SP_1241 PE = 1 SV = 1 - [Q97QH2_STRPN] Q97RS878.31 29 378 328.02 Lactate oxidase OS =- Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = lctO-2 PE = 4 SV= 1 - [Q97RS8_STRPN] Q97PW1 63.71 25 463 316.66 PeptidoglycanN-acetylglucosamine N- deacetylase A OS =- Streptococcus terminallypneumoniae serotype 4 (strain ATCC anchored BAA-334/TIGR4) GN = pgdA PE= 4 SV = 1 - (No CS) [Q97PW1_STRPN] Q97PT6 75.85 26 468 315.76 ATPsynthase subunit beta OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = atpD PE = 3 SV = 1 -[ATPB_STRPN] Q97NQ3 58.55 22 345 310 Uncharacterized protein OS = N-Streptococcus pneumoniae serotype 4 terminally (strain ATCCBAA-334/TIGR4) anchored GN = SP_1967 PE = 4 SV = 1 - (No CS)[Q97NQ3_STRPN] Q97R09 58.94 17 341 309.62 Iron-compound ABC transporter,iron Lipid compound-binding protein OS = anchored Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1032 PE = 1 SV= 1 - [Q97R09_STRPN] P18791 69.8 28 659 298.67 Oligopeptide-bindingprotein AmiA Lipid OS =- Streptococcus pneumoniae anchored serotype 4(strain ATCC BAA-334/ TIGR4) GN = amiA PE = 1 SV = 3 - [AMIA_STRPN]Q97RZ7 63.03 15 238 283.45 Uncharacterized protein OS = LipidStreptococcus pneumoniae serotype 4 anchored (strain ATCC BAA-334/TIGR4)GN = SP_0629 PE = 1 SV = 1- [Q97RZ7_STRPN] P0A4G2 65.05 18 309 281.58Manganese ABC transporter substrate- Lipid binding lipoprotein OS =-Streptococcus anchored pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4)GN = psaA PE = 1 SV = 1 - [MTSA_STRPN] P66112 60.5 10 119 280.56 50Sribosomal protein L20 OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = rplT PE = 3 SV = 1 -[RL20_STRPN] Q97PM5 63.3 23 376 278.8 Sugar ABC transporter, ATP-bindingIntracellular protein OS =- Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = msmK PE = 4 SV = 1 - [Q97PM5_STRPN] P0A4M741.57 23 498 271.04 Oligopeptide transport system Multi- permeaseprotein AmiC OS = transmembrane Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = amiC PE = 3 SV = 1 - [AMIC_STRPN]Q97SP4 48.18 12 303 262.56 PTS system, mannose-specific IID Multi-component OS =- Streptococcus transmembrane pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = SP_0282 PE = 4 SV = 1 - [Q97SP4_STRPN]Q97NL3 60.74 34 731 258.29 Penicillin-binding protein 2A OS = N-Streptococcus pneumoniae serotype 4 terminally (strain ATCCBAA-334/TIGR4) anchored GN = pbp2A PE = 4 SV = 1 - (No CS)[Q97NL3_STRPN] P14677 61.33 32 750 257.72 Penicillin-binding protein 2xOS = N- Streptococcus pneumoniae serotype 4 terminally (strain ATCCBAA-334/TIGR4) anchored GN = pbpX PE = 1 SV = 2 - [PBPX_STRPN] (No CS)Q97PT4 38.92 21 501 249.37 ATP synthase subunit alpha OS = Multi-Streptococcus pneumoniae serotype 4 transmembrane (strain ATCCBAA-334/TIGR4) GN = atpA PE = 3 SV = 1 - [ATPA_STRPN] Q97NB5 52.66 30621 236.54 Choline binding protein PcpA OS = Secretory Streptococcuspneumoniae serotype 4 (released) (strain ATCC BAA-334/TIGR4) (with CS)GN = pcpA PE = 1 SV = 1 - [Q97NB5_STRPN] Q97PQ2 33.26 21 914 236.23Cation-transporting ATPase, E1-E2 Multi- family OS =- Streptococcuspneumoniae transmembrane serotype 4 (strain ATCC BAA-334/ TIGR4) GN =SP_1551 PE = 3 SV = 1 - [Q97PQ2_STRPN] P0A475 50.36 8 137 229.6 50Sribosomal protein L16 OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = rplP PE = 1 SV = 1 -[RL16_STRPN] Q97PU3 64.03 23 278 227.7 Amino acid ABC transporter, aminoLipid acid-binding protein OS =- Streptococcus anchored pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = aatB PE = 4 SV = 1 -[Q97PU3_STRPN] Q97RC5 50.3 20 501 223.13 Pyruvate kinase OS =-Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = pyk PE = 3 SV = 1 - [Q97RC5_STRPN] Q97NS5 65.68 19338 221.3 Putative transcriptional regulator OS = SecretoryStreptococcus pneumoniae serotype 4 (released) (strain ATCCBAA-334/TIGR4) (with CS) GN = SP_1942 PE = 4 SV = 1 - [Q97NS5_STRPN]Q97RP4 75.22 14 230 213.93 Cell division ABC transporter, ATP-Intracellular binding protein FtsE OS =- Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = ftsE PE = 3 SV = 1 -[Q97RP4_STRPN] Q97RN2 66.67 13 267 207.19 Peptidyl-prolyl cis-transisomerase, Lipid cyclophilin-type OS =- Streptococcus anchoredpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0771 PE = 3 SV= 1 - [Q97RN2_STRPN] P66565 61.58 21 203 205.82 30S ribosomal protein S4OS = Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpsD PE = 3 SV = 1 - [RS4_STRPN] Q97QH1 61.79 10 246202.96 Amino acid ABC transporter, ATP- Intracellular binding protein OS=- Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_1242 PE = 3 SV = 1 - [Q97QH1_STRPN] I6L8N9 40.91 18 506 201.9 Cholinetransporter OS =- Streptococcus Multi- pneumoniae serotype 4 (strainATCC transmembrane BAA-334/TIGR4) GN = proWX PE = 3 SV = 1 -[I6L8N9_STRPN] Q97N69 55.22 15 335 201.71 Putative ABC transporter,substrate- Lipid binding protein OS =- Streptococcus anchored pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_2197 PE = 4 SV = 1 -[Q97N69_STRPN] P65887 53.97 23 428 199.01 Adenylosuccinate synthetase OS= Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = purA PE = 1 SV = 1 - [PURA_STRPN] P18766 60.71 13308 198.48 Oligopeptide transport ATP-binding Intracellular protein AmiFOS =- Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN= amiF PE = 3 SV = 2 - [AMIF_STRPN] Q97QB4 61.83 23 448 197.42 Glutamatedehydrogenase OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = gdhA PE = 3 SV = 1 - [Q97QB4_STRPN]Q97SP5 55.18 19 444 196.27 Aminopeptidase C OS =- StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =pepC PE = 4 SV = 1 - [Q97SP5_STRPN] Q97RP3 40.58 17 308 193.01 Celldivision protein FtsX OS = Multi- Streptococcus pneumoniae serotype 4transmembrane (strain ATCC BAA-334/TIGR4) GN = ftsX PE = 3 SV = 1 -[Q97RP3_STRPN] Q97T46 36.84 5 95 189.78 Putative bacteriocin OS =-Streptococcus Secretory pneumoniae serotype 4 (strain ATCC (released)BAA-334/TIGR4) GN = SP_0109 PE = 4 (no CS) SV = 1 - [Q97T46_STRPN]Q97PD6 51.85 31 837 188.69 Protein translocase subunit SecA 1 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = secA1 PE = 3 SV = 1 - [SECA1_STRPN] P0A4A7 60.58 13137 187.87 30S ribosomal protein S12 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rpsL PE = 3 SV =1 - [RS12_STRPN] P0A2U8 52.96 20 355 185.88 Oligopeptide transportATP-binding Intracellular protein AmiE OS =- Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = amiE PE = 3 SV = 1 -[AMIE_STRPN] Q97RQ0 70.21 18 386 182.7 Branched-chain amino acid ABCLipid transporter, amino acid-binding anchored protein OS =-Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =livJ PE = 4 SV = 1 - [Q97RQ0_STRPN] P0A3M9 67.68 16 328 176.61 L-lactatedehydrogenase OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = ldh PE = 3 SV = 2 - [LDH_STRPN] Q97RG949.51 20 511 174.4 Sugar ABC transporter, ATP-binding Intracellularprotein OS =- Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0846 PE = 3 SV = 1 - [Q97RG9_STRPN] P0A3M5 43.3822 680 166.84 Penicillin-binding protein 2B OS = N- Streptococcuspneumoniae serotype 4 terminally (strain ATCC BAA-334/TIGR4) anchored GN= penA PE = 3 SV = 1 - [PBP2_STRPN] (No CS) Q97N53 63.24 12 272 163.14Cell shape-determining protein MreC N- OS =- Streptococcus pneumoniaeterminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = mreC PE= 3 SV = 1 - (No CS) [Q97N53_STRPN] P95829 49.26 20 607 161.66 Chaperoneprotein Dnak OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = dnaK PE = 3 SV = 2 - [DNAK_STRPN]Q97S19 67.89 7 109 156.35 Cell cycle protein GpsB OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =gpsB PE = 3 SV = 1 - [GPSB_STRPN] Q97NE4 36.3 26 821 153.09Penicillin-binding protein 1B OS = N- Streptococcus pneumoniae serotype4 terminally (strain ATCC BAA-334/TIGR4) anchored GN = pbp1B PE = 4 SV =1 - (No CS) [Q97NE4_STRPN] I6L8W3 71.49 15 242 152.01 Cholinetransporter OS =- Streptococcus Intracellular pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = proV PE = 3 SV = 1 - [I6L8W3_STRPN]P0A335 52.04 21 540 151.44 60 kDa chaperonin OS =- StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =groL PE = 1 SV = 1 - [CH60_STRPN] Q97NG0 44.77 17 449 145.04Glucose-6-phosphate isomerase OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = pgi PE = 3 SV =1 - [G6PI_STRPN] Q97Q31 57.53 12 292 143.85 Phosphate-binding proteinPstS 1 OS = Lipid Streptococcus pneumoniae serotype 4 anchored (strainATCC BAA-334/TIGR4) GN = pstS1 PE = 1 SV = 1 - [PSTS1_STRPN] Q97RW951.81 16 332 139.95 Putative pneumococcal surface protein Lipid OS =-Streptococcus pneumoniae anchored serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0667 PE = 1 SV = 1 - [Q97RW9_STRPN] P22976 44.98 18 658139.63 Probable transketolase OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = tkt PE = 3 SV =2 - [TKT_STRPN] Q97SQ9 42.52 17 602 138.65Glutamine--fructose-6-phosphate Intracellular aminotransferase[isomerizing] OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = glmS PE = 3 SV = 3 - [GLMS_STRPN] Q97PA9 33.84 18659 132.79 Serine/threonine-protein kinase StkP Intracellular/ OS =-Streptococcus pneumoniae TMH start serotype 4 (strain ATCC BAA-334/AFTER 60 TIGR4) GN = stkP PE = 1 SV = 1 - [STKP2_STRPN] Q97Q34 70.41 18267 131.91 Phosphate import ATP-binding protein Intracellular PstB 2 OS=- Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =pstB2 PE = 3 SV = 1 - [PSTB2_STRPN] Q97QX5 55.81 14 344 130.66Uncharacterized protein OS = Secretory( Streptococcus pneumoniaeserotype 4 released) (strain ATCC BAA-334/TIGR4) (with CS) GN = SP_1069PE = 1 SV = 1 - [Q97QX5_STRPN] I6L8V0 55.84 18 351 130.62 Capsularpolysaccharide biosynthesis Intracellular protein Cps4J OS =-Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =cap4J PE = 4 SV = 1 - [I6L8V0_STRPN] Q97SD1 43.38 17 521 119.96 Aminoacid ABC transporter, amino Multi- acid-binding protein/permease proteintransmembrane OS =- Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_0453 PE = 4 SV = 1 - [Q97SD1_STRPN] Q97SR1 50.7321 617 119.02 Proline--tRNA ligase OS =- Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = proS PE = 3 SV =1 - [SYP_STRPN] P0A4D7 36.07 14 524 117.93 DEAD-box ATP-dependent RNAIntracellular helicase CshA OS =- Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = cshA PE = 3 SV = 1 - [CSHA_STRPN]Q97P40 68.81 12 202 117.92 Putative general stress protein 24 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1804 PE = 4 SV = 1 - [Q97P40_STRPN] Q97NK0 41 21883 117.4 Aldehyde-alcohol dehydrogenase OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_2026 PE = 3 SV = 1 - [Q97NK0_STRPN] Q97N99 51.82 14 274 115.56 SPFHdomain/Band 7 family OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_2156 PE = 4 SV = 1 -[Q97N99_STRPN] Q9L7Q2 17.63 23 190 6 114.8 Zinc metalloprotease ZmpB OS= Multi- Streptococcus pneumoniae serotype 4 transmembrane (strain ATCCBAA-334/TIGR4) GN = zmpB PE = 3 SV = 2 - [ZMPB_STRPN] P67293 43.9 4 82114.56 UPF0154 protein SP_1882 OS = N- Streptococcus pneumoniae serotype4 terminally (strain ATCC BAA-334/TIGR4) anchored GN = SP_1882 PE = 3 SV= 1 - (No CS) [Y1882_STRPN] Q97N37 61.83 14 393 114.16 Serine proteaseOS =- Streptococcus N- pneumoniae serotype 4 (strain ATCC terminallyBAA-334/TIGR4) GN = SP_2239 PE = 1 anchored SV = 1 - [Q97N37_STRPN] (NoCS) P66359 55.12 8 127 112.68 30S ribosomal protein S11 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpsK PE = 3 SV = 1 - [RS11_STRPN] Q97PX1 30.72 12459 110.91 NADH oxidase OS =- Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = nox PE = 4 SV = 1 -[Q97PX1_STRPN] P35592 43.64 21 660 109.44 Oligopeptide-binding proteinAliA OS = Lipid Streptococcus pneumoniae serotype 4 anchored (strainATCC BAA-334/TIGR4) GN = aliA PE = 3 SV = 4 - [ALIA_STRPN] Q97PC4 28.6611 492 108.41 ABC transporter, ATP-binding protein Multi- OS =-Streptococcus pneumoniae transmembrane serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1715 PE = 4 SV = 1 - [Q97PC4_STRPN] Q97P68 69.23 8 104104.87 Thioredoxin OS =- Streptococcus Intracellular pneumoniae serotype4 (strain ATCC BAA-334/TIGR4) GN = trx PE = 3 SV = 1 - [Q97P68_STRPN]Q97PL8 41.55 12 438 101 Oxidoreductase, pyridine nucleotide-Intracellular disulfide, class I OS =- Streptococcus pneumoniae serotype4 (strain ATCC BAA-334/TIGR4) GN = SP_1588 PE = 3 SV = 1 -[Q97PL8_STRPN] Q975X2 68.78 17 189 97.17 Uncharacterized protein OS =Lipid Streptococcus pneumoniae serotype 4 anchored (strain ATCCBAA-334/TIGR4) GN = SP_0191 PE = 1 SV = 1 - [Q97SX2_STRPN] P63413 43.9414 396 96.43 Acetate kinase OS =- Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = ackA PE = 3 SV = 1 -[ACKA_STRPN] Q97SJ6 52.61 11 230 96.34 Capsular polysaccharidebiosynthesis Multi- protein CpsC OS =- Streptococcus transmembranepneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = cpsC PE = 3 SV =1 - [CPSC_STRPN] Q97Q67 38.08 11 344 95.87 Conserved domain protein OS =Multi- Streptococcus pneumoniae serotype 4 transmembrane (strain ATCCBAA-334/TIGR4) GN = SP_1363 PE = 4 SV = 1 - [Q97Q67_STRPN] P0A4S1 49.1510 293 93.48 Fructose-bisphosphate aldolase OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = fbaPE = 3 SV = 1 - [ALF_STRPN] Q97SR2 37.71 11 419 93.31 Putative zincmetalloprotease SP_0263 Multi- OS =- Streptococcus pneumoniaetransmembrane serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_0263 PE =3 SV = 1 - [Y263_STRPN] Q97NP5 25.65 9 308 92.53 Membrane proteininsertase YidC 1 Multi- OS =- Streptococcus pneumoniae transmembraneserotype 4 (strain ATCC BAA-334/ TIGR4) GN = yidC1 PE = 3 SV = 1 -[YIDC1_STRPN] Q97SJ4 44.44 15 360 91.77 Uncharacterized protein OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0355 PE = 4 SV = 1 - [Q97SJ4_STRPN] Q97RK0 19.8310 575 91.07 Septation ring formation regulator N- EzrA OS = =-Streptococcus pneumoniae terminally serotype 4 (strain ATCC BAA-334/anchored TIGR4) GN = ezrA PE = 3 SV = 1 - (No CS) [EZRA_STRPN] P6337371.43 11 252 91.01 Phosphate import ATP-binding protein IntracellularPstB 1 OS =- Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = pstB1 PE = 3 SV = 1 - [PSTB1_STRPN] Q97S89 51.01 13 39889.47 Phosphoglycerate kinase OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = pgk PE = 3 SV =1 - [PGK_STRPN] Q97PT5 47.26 12 292 85.91 ATP synthase gamma chain OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = atpG PE = 3 SV = 1 - [ATPG_STRPN] Q97514 32.65 9 34085.66 Choline binding protein C OS = N- Streptococcus pneumoniaeserotype 4 terminally (strain ATCC BAA-334/TIGR4) anchored GN = cbpC PE= 4 SV = 1 - [Q97SI4_STRPN] (with CS) I6L858 39.71 14 481 85.62 Capsularpolysaccharide biosynthesis Multi- protein Cps4A OS =- Streptococcustransmembrane pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =cps4A PE = 4 SV = 1 - [I6L8S8_STRPN] Q97PY0 65.59 13 247 85.07 Aminoacid ABC transporter, ATP- Intracellular binding protein OS =-Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_1460 PE = 3 SV = 1 - [Q97PYO_STRPN] Q97SQ4 46.79 9 156 83.39 30Sribosomal protein S7 OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = rpsG PE = 3 SV = 1 -[RS7_STRPN] P0A3Y3 60 10 230 82.54 2,3-bisphosphoglycerate-dependentIntracellular phosphoglycerate mutase OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = gpmA PE = 3 SV = 1 -[GPMA_STRPN] P0A4C3 59.91 11 217 82.05 30S ribosomal protein S3 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpsC PE = 3 SV = 1 - [RS3_STRPN] P65607 38.46 11 33879.74 Ornithine carbamoyltransferase, Intracellular catabolic OS =-Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =arcB PE = 3 SV = 1 - [OTCC_STRPN] O05703 39.92 12 501 79.68 Zinc-bindinglipoprotein AdcA OS = Lipid Streptococcus pneumoniae serotype 4 anchored(strain ATCC BAA-334/TIGR4) GN = adcA PE = 3 SV = 4 - [ADCA_STRPN]I6L8U0 27.16 11 394 79.39 UDP-N-acetylglucosamine 2-epimeraseIntracellular OS =- Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = cps4L PE = 3 SV = 1 - [I6L8U0_STRPN] Q97SI7 28.23 8464 78.19 Mid-cell-anchored protein Z OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = mapZ PE = 1 SV =1 - [Q97SI7_STRPN] Q97N56 58.3 15 259 78.0 30S ribosomal protein S2 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpsB PE = 3 SV = 1 - [RS2_STRPN] Q97SI6 35.44 13 47477.15 6-phosphogluconate dehydrogenase, Intracellular decarboxylating OS=- Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =gnd PE = 3 SV = 1 - [Q97SI6_STRPN] Q97PG0 27.1 7 262 77.07 Cell divisionprotein DivIVA OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = divIVA PE = 4 SV = 1 - [Q97PG0_STRPN]Q97RF9 32 11 400 76.46 Ribosomal protein S1 OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =rpsA PE = 4 SV = 1 - [Q97RF9_STRPN] Q97T80 15.68 17 1856 76.28 Zincmetalloprotease ZmpC OS = LPXTG Streptococcus pneumoniae serotype 4Cell-wall (strain ATCC BAA-334/TIGR4) anchored GN = zmpC PE = 3 SV = 1 -[ZMPC_STRPN] P0A2Z2 62.8 9 164 75.56 ATP synthase subunit b OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = atpF PE = 3 SV = 1 - [ATPF_STRPN] I6L859 47.7 10 41373.22 D-alanyl-D-alanine carboxypeptidase N- OS =- Streptococcuspneumoniae terminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4)GN = dacA PE = 3 SV = 1 - (with CS) [I6L859_STRPN] Q97PU4 80 7 115 71.8Bacterocin transport accessory protein Intracellular OS =- Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = bta PE = 4 SV =1 - [Q97PU4_STRPN] Q97SU6 70.34 7 118 71.64 50S ribosomal protein L18 OS= Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rplR PE = 3 SV = 1 - [RL18_STRPN] P63742 35.46 11502 71.21 Glycerol kinase OS =- Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = glpK PE = 3 SV = 1 -[GLPK_STRPN] Q97SN4 32.31 6 130 68.89 30S ribosomal protein S9 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpsI PE = 3 SV = 1 - [RS9_STRPN] P66907 31.84 6 38068.13 Queuine tRNA-ribosyltransferase OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = tgt PE = 3 SV =1 - [TGT_STRPN] Q97SF8 32.16 12 454 65.06 Aspartokinase OS =-Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0413 PE = 3 SV = 1 - [Q97SF8_STRPN] P0A471 82.799 122 62.07 50S ribosomal protein L7/L12 OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =rplL PE = 1 SV = 2 - [RL7_STRPN] I6L8W8 21.08 8 332 61.96 Cholinebinding protein J OS = N- Streptococcus pneumoniae serotype 4 terminally(strain ATCC BAA-334/TIGR4) anchored GN = cbpJ PE = 4 SV = 1 -[I6L8W8_STRPN] (with CS) P0A4J6 40.8 5 201 61.71 Superoxide dismutase[Mn] OS = Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = sodA PE = 1 SV = 2 - [SODM_STRPN] Q97T52 30.03 14616 60.5 Putative capsular polysaccharide Multi- biosynthesis protein OS=- Streptococcus transmembrane pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0103 PE = 4 SV = 1 - [Q97T52_STRPN] Q97RE5 17.086 650 60.23 PTS system, fructose specific IIABC Multi- components OS =-Streptococcus transmembrane pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0877 PE = 4 SV = 1 - [Q97RE5_STRPN] Q97SV4 54.119 207 60.13 50S ribosomal protein L4 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rplD PE = 3 SV =1 - [RL4_STRPN] Q97RK7 19.46 12 848 59.83 Aminopeptidase N OS =-Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = pepN PE = 4 SV = 1 - [Q97RK7_STRPN] Q97Q62 34.67 11424 59.21 Psr protein OS =- Streptococcus Intracellular/ pneumoniaeserotype 4 (strain ATCC TMH start BAA-334/TIGR4) GN = SP_1368 PE = 4AFTER 60 SV = 1 - [Q97Q62_STRPN] O07344 26.96 5 204 58.64 Signalpeptidase I OS =- Streptococcus N- pneumoniae serotype 4 (strain ATCCterminally BAA-334/TIGR4) GN = lepB PE = 3 SV = 2 - anchored [LEP_STRPN](No CS) P0A3S3 34.67 6 274 57.63 DNA-entry nuclease OS =- StreptococcusN- pneumoniae serotype 4 (strain ATCC terminally BAA-334/TIGR4) GN =endA PE = 1 SV = 1 - anchored [NUCE_STRPN] (No CS) Q97PU5 29.55 10 57257.38 Phosphoglucomutase OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = pgm PE = 3 SV = 1 -[Q97PU5_STRPN] Q97RC6 31.34 8 335 57.3 ATP-dependent 6- Intracellularphosphofructokinase OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = pfkA PE = 3 SV = 1 - [PFKA_STRPN] Q97SV5 37.5 6208 56.96 50S ribosomal protein L3 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rplC PE = 3 SV =1 - [RL3_STRPN] Q97RL9 25.56 6 399 56.21 Uncharacterized protein OS = N-Streptococcus pneumoniae serotype 4 terminally (strain ATCCBAA-334/TIGR4) anchored GN = SP_0785 PE = 4 SV = 1 - (No CS)[Q97RL9_STRPN] P66278 18.18 2 66 55.84 50S ribosomal protein L35 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpmI PE = 3 SV = 1 - [RL35_STRPN] I6L8N0 40.82 12365 55.32 UDP-N-acetylglucosamine-2-epimerase Intracellular OS =-Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =cps4I PE = 3 SV = 1 - [I6L8N0_STRPN] Q97PQ8 57.34 6 143 54.74Uncharacterized protein OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1545 PE = 4 SV = 1 -[Q97PQ8_STRPN] Q97TA6 33.64 10 330 52.71 Phosphate acyltransferase OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = plsX PE = 3 SV = 1 - [PLSX_STRPN] Q97PK5 27.22 2 15852.32 Uncharacterized protein OS = N- Streptococcus pneumoniae serotype4 terminally (strain ATCC BAA-334/TIGR4) anchored GN = SP_1604 PE = 4 SV= 1 - (No CS) [Q97PK5_STRPN] Q97SN5 67.57 8 148 51.12 50S ribosomalprotein L13 OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = rplM PE = 3 SV = 1 - [Q97SN5_STRPN]Q97RV8 60.32 8 126 49.28 Uncharacterized protein OS = N- Streptococcuspneumoniae serotype 4 terminally (strain ATCC BAA-334/TIGR4) anchored GN= SP_0678 PE = 4 SV = 1 - (No CS) [Q97RV8_STRPN] Q97R14 30.25 7 324 48.6Uncharacterized protein OS = Secretory Streptococcus pneumoniae serotype4 (released) (strain ATCC BAA-334/TIGR4) (with CS) GN = SP_1027 PE = 4SV = 1 - [Q97R14_STRPN] P35597 15.04 7 778 48.34 Probablecation-transporting ATPase Multi- exp7 OS =- Streptococcus pneumoniaetransmembrane serotype 4 (strain ATCC BAA-334/ TIGR4) GN = exp7 PE = 3SV = 2 - [EXP7_STRPN] P65239 41.61 9 322 48.08 Ribose-phosphatepyrophosphokinase Intracellular 1 OS =- Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = prs1 PE = 3 SV = 1 -[KPRS1_STRPN] Q97Q37 46.13 7 271 47.67 Amino acid ABC transporter, aminoLipid acid-binding protein OS =- Streptococcus anchored pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1394 PE = 4 SV = 1 -[Q97Q37_STRPN] Q97R36 52.43 8 185 47.65 Thioredoxin family protein OS =Lipid Streptococcus pneumoniae serotype 4 anchored (strain ATCCBAA-334/TIGR4) GN = SP_1000 PE = 1 SV = 1 - [Q97R36_STRPN] Q97N48 26.815 276 45.87 Uncharacterized protein OS = Intracellular/ Streptococcuspneumoniae serotype 4 TMH start (strain ATCC BAA-334/TIGR4) AFTER 60 GN= SP_2223 PE = 4 SV = 1 - [Q97N48_STRPN] I6L8R1 22.79 4 215 45.06 ABCtransporter, ATP-binding protein Intracellular Vexp2 OS =- Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = vex2 PE = 3 SV =1 - [I6L8R1_STRPN] Q97SC6 20.93 10 774 44.93 Formate acetyltransferaseOS = Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = pfl PE = 4 SV = 1 - [Q97SC6_STRPN] Q97QD5 51.08 8186 44.75 LemA protein OS =- Streptococcus N- pneumoniae serotype 4(strain ATCC terminally BAA-334/TIGR4) GN = lemA PE = 1 SV = 1 -anchored [Q97QD5_STRPN] (No CS) Q97PF9 37.71 9 419 44.71 Cell divisionprotein FtsZ OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = ftsZ PE = 3 SV = 1 - [Q97PF9_STRPN]P0A3B7 36.71 7 346 44.56 Elongation factor Ts OS =- StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = tsfPE = 1 SV = 2 - [EFTS_STRPN] P72524 20.56 10 822 44.46 DNA gyrasesubunit A OS = Intracellular Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = gyrA PE = 3 SV = 3 - [GYRA_STRPN] P67282 26.2211 534 43.28 Ribonuclease Y OS =- Streptococcus N- pneumoniae serotype 4(strain ATCC terminally BAA-334/TIGR4) GN = rny PE = 3 SV = 1 - anchored[RNY_STRPN] (No CS) P66419 38.2 6 89 42.98 30S ribosomal protein S14 OS= Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpsN PE = 3 SV = 1 - [RS14_STRPN] Q97SV1 48.89 6 18042.64 50S ribosomal protein L5 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rplE PE = 3 SV =1 - [RL5_STRPN] Q97NW9 7.48 5 655 42.08 Trehalose PTS system, IIABCMulti- components OS =- Streptococcus transmembrane pneumoniae serotype4 (strain ATCC BAA-334/TIGR4) GN = SP_1884 PE = 4 SV = 1 -[Q97NW9_STRPN] Q97Q18 91.06 6 123 40.86 Uncharacterized protein OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1218 PE = 4 SV = 1 - [Q97Q18_STRPN] Q97T04 11.922 193 39.59 Uncharacterized protein OS = Multi- Streptococcus pneumoniaeserotype 4 transmembrane (strain ATCC BAA-334/TIGR4) GN = SP_0157 PE = 4SV = 1 - [Q97T04_STRPN] Q97PB8 9.09 4 627 39.49 PTS system IIABCcomponents OS = Multi- Streptococcus pneumoniae serotype 4 transmembrane(strain ATCC BAA-334/TIGR4) GN = SP_1722 PE = 4 SV = 1 - [Q97PB8_STRPN]Q97P19 57.3 5 89 39.48 30S ribosomal protein S15 OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =rpsO PE = 3 SV = 1 - [RS15_STRPN] P61182 40.35 4 114 39.35 50S ribosomalprotein L22 OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = rplV PE = 1 SV = 1 - [RL22_STRPN]Q97SU7 51.69 6 178 38.14 50S ribosomal protein L6 OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =rplF PE = 3 SV = 1 - [RL6_STRPN] Q97NB9 39.1 8 335 38.04 Uncharacterizedprotein OS = Multi- Streptococcus pneumoniae serotype 4 transmembrane(strain ATCC BAA-334/TIGR4) GN = SP_2132 PE = 4 SV = 1 - [Q97NB9_STRPN]Q97QP7 5.34 8 2004 37.84 Immunoglobulin A1 protease OS = Multi-Streptococcus pneumoniae serotype 4 transmembrane (strain ATCCBAA-334/TIGR4) GN = iga PE = 3 SV = 1 - [IGA1A_STRPN] Q97N55 23.72 6 39237.16 Secreted 45 kd protein OS = Secretory Streptococcus pneumoniaeserotype 4 (released) (strain ATCC BAA-334/TIGR4) (with CS) GN = usp45PE = 4 SV = 1 - [Q97N55_STRPN] P66942 18.25 4 252 36.63 Triosephosphateisomerase OS = Intracellular Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = tpiA PE = 3 SV = 1 - [TPIS_STRPN] Q97QC6 66.097 115 35.81 50S ribosomal protein L19 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rplS PE = 3 SV =1 - [RL19_STRPN] I6L8S7 14.6 5 459 35.54 Transmembrane protein Vexp3 OS= Multi- Streptococcus pneumoniae serotype 4 transmembrane (strain ATCCBAA-334/TIGR4) GN = vex3 PE = 4 SV = 1 - [I6L8S7_STRPN] P35595 12.4 6726 35.44 PTS system glucose-specific EIICBA Multi- component OS =-Streptococcus transmembrane pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = exp5 PE = 3 SV = 2 - [PTG3C_STRPN] Q97PV1 14.88 3289 34.89 Putative glycerol uptake facilitator Multi- protein OS =-Streptococcus transmembrane pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1491 PE = 3 SV = 1 - [Q97PV1_STRPN] Q97NM6 21.046 404 33.79 Aminotransferase, class I OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1994 PE = 4 SV= 1 - [Q97NM6_STRPN] Q97SD2 45.93 5 246 33.44 Amino acid ABCtransporter, ATP- Intracellular binding protein OS =- Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0452 PE = 3 SV= 1 - [Q97SD2_STRPN] Q97NN3 36.15 6 213 33.1 ABC transporter,ATP-binding protein Intracellular OS =- Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1987 PE = 1 SV = 1 -[Q97NN3_STRPN] Q97RJ0 23.77 5 244 33.09 Amino acid ABC transporter, ATP-Intracellular binding protein OS =- Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = SP_0824 PE = 3 SV = 1 - [Q97RJ0_STRPN]Q97NJ7 27.27 3 99 32.76 Preprotein translocase, YajC subunit N- OS =-Streptococcus pneumoniae terminally serotype 4 (strain ATCC BAA-334/anchored TIGR4) GN = yajC-2 PE = 4 SV = 1 - (No CS) [Q97NJ7_STRPN]P66095 34.5 8 229 32.4 50S ribosomal protein L1 OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =rplA PE = 3 SV = 1 - [RL1_STRPN] Q97NT7 38.61 6 158 32.3 Uncharacterizedprotein OS = Multi- Streptococcus pneumoniae serotype 4 transmembrane(strain ATCC BAA-334/TIGR4) GN = SP_1926 PE = 4 SV = 1 - [Q97NT7_STRPN]P63667 29.5 4 139 31.7 ATP synthase epsilon chain OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =atpC PE = 3 SV = 1 - [ATPE_STRPN] Q97T58 18.08 4 354 31.6 Conserveddomain protein Multi- OS = Streptococcus pneumoniae transmembraneserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_0097 PE = 4 SV = 1 -[Q97T58_STRPN] Q97R06 36.74 6 264 31.17 Iron-compound ABC transporter,ATP- Intracellular binding protein OS =- Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1035 PE = 3 SV = 1 -[Q97R06_STRPN] Q97PQ5 20.34 6 531 30.81 Uncharacterized protein OS =Multi- Streptococcus pneumoniae serotype 4 transmembrane (strain ATCCBAA-334/TIGR4) GN = SP_1548 PE = 4 SV = 1 - [Q97PQ5_STRPN] Q97R44 33.912 115 30.62 Uncharacterized protein OS = Intracellular/ Streptococcuspneumoniae serotype 4 TMH start (strain ATCC BAA-334/TIGR4) AFTER 60 GN= SP_0990 PE = 4 SV = 1 - [Q97R44_STRPN] Q97SW3 47.73 3 88 29.98Uncharacterized protein OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0201 PE = 4 SV = 1 -[Q97SW3_STRPN] I6L8N6 27.01 4 211 29.5 Capsular polysaccharidebiosynthesis N- protein Cps4E OS =- Streptococcus terminally pneumoniaeserotype 4 (strain ATCC anchored BAA-334/TIGR4) GN = cps4E PE = 4 SV =1 - (No CS) [I6L8N6_STRPN] Q97SU3 34.25 5 146 28.53 50S ribosomalprotein L15 OS = Intracellular Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = rplO PE = 3 SV = 1 - [RL15_STRPN]Q97QS6 25.86 6 379 28.15 Glycogen biosynthesis protein GlgDIntracellular OS =- Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = glgD PE = 4 SV = 1 - [Q97QS6_STRPN] Q97P39 52.24 267 27.23 UPF0337 protein SP_1805 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1805 PE = 3 SV= 1 - [Y1805_STRPN] P06653 20.44 6 318 25.81 Autolysin OS =-Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = lytA PE = 1 SV = 2 - [ALYS_STRPN] Q97PX9 12.78 5 26625.41 Amino acid ABC transporter, permease Multi- protein OS =-Streptococcus transmembrane pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1461 PE = 3 SV = 1 - [Q97PX9_STRPN] Q97QW4 17.655 425 24.84 Uncharacterized protein OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1083 PE = 4 SV= 1 - [Q97QW4_STRPN] Q97S93 13.27 7 535 23.79 CTP synthase OS =-Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = pyrG PE = 3 SV = 1 - [PYRG_STRPN] Q97SX1 26.14 2 8822 UPF0297 protein SP_0192 OS = Intracellular Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0192 PE = 1 SV = 1 -[Y192_STRPN] Q97RH7 50 4 78 21.34 30S ribosomal protein S20 OS =Intracellular Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rpsT PE = 3 SV = 1 - [RS20_STRPN] P66050 45.78 5 16620.98 50S ribosomal protein L10 OS = Intracellular Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rplJ PE = 3 SV =2 - [RL10_STRPN] Q97RW6 32.14 3 56 20.85 Uncharacterized protein OS =Secretory Streptococcus pneumoniae serotype 4 (released) (strain ATCCBAA-334/TIGR4) (with CS) GN = SP_0670 PE = 4 SV = 1 - [Q97RW6_STRPN]Q9FBB7 29.41 4 255 20.65 Acetyl-coenzyme A carboxylase Intracellularcarboxyl transferase subunit alpha OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = accA PE = 1 SV = 1 -[ACCA_STRPN] I6L8V8 30.45 4 243 20.58 3-oxoacyl-[acyl-carrier protein]Intracellular reductase OS =- Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = fabG PE = 3 SV = 1 - [I6L8V8_STRPN]P66581 41.46 4 164 20.46 30S ribosomal protein S5 OS = IntracellularStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =rpsE PE = 3 SV = 1 - [RS5_STRPN] Q97T45 11.82 5 694 20.39Uncharacterized protein OS = Multi- Streptococcus pneumoniae serotype 4transmembrane (strain ATCC BAA-334/TIGR4) GN = SP_0110 PE = 4 SV = 1 -[Q97T45_STRPN] Q97P18 18.31 4 355 20.06 ABC transporter,substrate-binding Lipid protein OS =- Streptococcus anchored pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1826 PE = 4 SV = 1 -[Q97P18_STRPN]

TABLE 5 Proteins identified in MV_(L) from serotype 3. The accessionnumbers refer to SwissProt on the date of filing. Accession # Coverage #Peptides #AAs Score Description Localization B2IQV2 91.09 50 404 5251.94Elongation factor Tu OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = tuf PE = 3 SV = 1 - [B2IQV2_STRPS] B2IPX8 95.62 40 4342291.84 Enolase OS =- Streptococcus Intracellular pneumoniae (strainCGSP14) GN = eno PE = 3 SV = 1 - [ENO_STRPS] B2INW2 70.86 42 374 2221.7Lipoprotein OS =- Streptococcus Lipid pneumoniae (strain CGSP14)anchored GN = SPCG_0787 PE = 4 SV = 1 - [B2INW2_STRPS] B2ISJ9 85.28 67693 1851.66 Elongation factor G OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = fusA PE = 3 SV = 1 - [EFG_STRPS] B2IMI773.82 33 359 1675.28 Glyceraldehyde-3-phosphate Intracellulardehydrogenase OS =- Streptococcus pneumoniae (strain CGSP14) GN = gapAPE = 3 SV = 1 - [B2IMI7_STRPS] B2IM39 82.94 101 1225 1348.46DNA-directed RNA polymerase Intracellular subunit beta' OS =-Streptococcus pneumoniae (strain CGSP14) GN = rpoC PE = 3 SV = 1 -[RPOC_STRPS] B2IMJ9 86.85 68 890 1321.16 Aldehyde-alcohol dehydrogenaseIntracellular OS =- Streptococcus pneumoniae (strain CGSP14) GN = adhEPE = 3 SV = 1 - [B2IMJ9_STRPS] B2IPD4 73.16 38 313 1171.98 Foldaseprotein PrsA OS = Lipid Streptococcus pneumoniae (strain anchoredCGSP14) GN = prsA PE = 3 SV = 1 - [PRSA_STRPS] B2INY1 91 53 400 1141.8430S ribosomal protein S1 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = rpsA PE = 4 SV = 1 - [B2INY1_STRPS] B2IP53 87.46 27335 830.14 ATP-dependent 6- Intracellular phosphofructokinase OS =Streptococcus pneumoniae (strain CGSP14) GN = pfkA PE = 3 SV = 1 -[PFKA_STRPS] B2IQX0 90.6 38 468 772.66 ATP synthase subunit beta OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = atpD PE = 3SV = 1 - [ATPB_STRPS] B2IRV7 72.76 62 837 760.12 Protein translocasesubunit SecA Intracellular OS =- Streptococcus pneumoniae (strainCGSP14) GN = secA PE = 3 SV = 1 - [B2IRV7_STRPS] B2IN25 78.22 51 450751.01 Maltose/maltodextrin ABC Lipid transporter, maltose/maltodextrin-anchored binding protein OS =- Streptococcus pneumoniae (strain CGSP14)GN = SPCG_2072 PE = 4 SV = 1 - [B2IN25_STRPS] B2INB2 72.76 41 591 749.62Pyruvate oxidase OS =- Streptococcus Intracellular pneumoniae (strainCGSP14) GN = spxB PE = 3 SV = 1 - [B2INB2_STRPS] B2ISZ0 69.04 47 659746.61 Oligopeptide ABC transporter, Lipid oligopeptide-binding proteinAmiA anchored OS =- Streptococcus pneumoniae (strain CGSP14) GN = amiAPE = 4 SV = 1 - [B2ISZ0_STRPS] B2ISK8 56.44 22 303 734.11 PTS system,mannose-specific IID Multi- component OS =- Streptococcus transmembranepneumoniae (strain CGSP14) GN = manN PE = 4 SV = 1 - [B2ISK8_STRPS]B2INN2 85.33 30 259 682.23 30S ribosomal protein S2 OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = rpsB PE = 3 SV = 2 -[RS2_STRPS] B2IP54 76.05 39 501 676.55 Pyruvate kinase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = SPCG_0873 PE= 3 SV = 1 - [B2IP54_STRPS] B2IR47 69.48 50 652 673.91 Oligopeptide ABCtransporter, Lipid oligopeptide-binding protein AliB anchored OS =-Streptococcus pneumoniae (strain CGSP14) GN = aliB PE = 4 SV = 1 -[B2IR47_STRPS] B2IR00 66.26 44 652 636.95 ATP-dependent zinc Multi-metalloprotease FtsH OS = transmembrane Streptococcus pneumoniae (strainCGSP14) GN = ftsH PE = 3 SV = 1 - [B2IR00_STRPS] B2IM40 77.67 50 824633.52 DNA-directed RNA polymerase Intracellular subunit beta OS =-Streptococcus pneumoniae (strain CGSP14) GN = rpoB PE = 3 SV = 1 -[B2IM40_STRPS] B2IS43 70.76 23 277 575.93 50S ribosomal protein L2 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rplB PE = 3SV = 1 - [RL2_STRPS] B2IS46 79.26 27 217 569.56 30S ribosomal protein S3OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpsC PE= 3 SV = 1 - [RS3_STRPS] B2IME4 64.95 51 930 553.42 Translationinitiation factor IF-2 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = infB PE = 3 SV = 1 - [IF2_STRPS] B2IQY1 73.14 43551 550.86 Uncharacterized protein OS = Intracellular/ Streptococcuspneumoniae (strain TMH start CGSP14) GN = SPCG_1503 PE = 4 SV = 1 -AFTER 60 [B2IQY1_STRPS] B2INT1 58.9 34 584 531.45 Septation ringformation regulator N- EzrA OS =- Streptococcus pneumoniae terminally(strain CGSP14) GN = ezrA PE = 3 SV = 1 - anchored [B2INT1_STRPS] (NoCS) B2IPY8 67.93 32 474 511.28 Glyceraldehyde-3-phosphate Intracellulardehydrogenase, NADP-dependent OS =- Streptococcus pneumoniae (strainCGSP14) GN = gapN PE = 3 SV = 1 - [B2IPY8_STRPS] B2IRQ6 79.23 20 313461.56 Manganese ABC transporter, Lipid manganese-binding adhesionanchored liprotein OS =- Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1623 PE = 3 SV = 1 - [B2IRQ6_STRPS] B2IPU8 63.14 36 719 457.32Ribonucleoside-diphosphate Intracellular reductase OS =- Streptococcuspneumoniae (strain CGSP14) GN = nrdE PE = 3 SV = 1 - [B2IPU8_STRPS]B2IM49 72.42 27 330 449.87 Aspartate--ammonia ligase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = asnA PE = 3 SV = 1 -[ASNA_STRPS] B2IQX2 68.86 31 501 447.54 ATP synthase subunit alpha OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = atpA PE = 3SV = 1 - [ATPA_STRPS] B2IMK3 70.36 32 658 435.4 Transketolase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = tktA PE = 3SV = 1 - [B2IMK3_STRPS] B2IRH9 64.45 36 616 434.01 Capsularpolysaccharide Multi- biosynthesis protein, putative OS = transmembraneStreptococcus pneumoniae (strain CGSP14) GN = SPCG_0098 PE = 4 SV = 1 -[B2IRH9_STRPS] B2IN95 82.06 37 496 432.16 Lysine--tRNA ligase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = lysS PE = 3SV = 1 - [SYK_STRPS] B2IS90 71.32 40 537 413.57 Ribonuclease Y OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = rny PE = 3SV = 1 - [B2IS90_STRPS] B2IRT9 66.06 32 442 412.96 Sugar ABCtransporter, sugar- Lipid binding protein OS =- Streptococcus anchoredpneumoniae (strain CGSP14) GN = SPCG_1656 PE = 4 SV = 1 - [B2IRT9_STRPS]B2IN10 73.37 15 338 401 Glycerol-3-phosphate Intracellular dehydrogenase[NAD(P)+] OS = Streptococcus pneumoniae (strain CGSP14) GN = gpsA PE = 3SV = 1 - [GPDA_STRPS] B2IP07 38.46 23 650 398.03 PTS system, fructosespecific IIABC Multi- components OS =- Streptococcus transmembranepneumoniae (strain CGSP14) GN = fruA PE = 4 SV = 1 - [B2IP07_STRPS]B2IPB5 52.94 10 119 388.03 50S ribosomal protein L20 OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = rplT PE = 3 SV = 1 -[RL20_STRPS] B2IS19 66.29 35 617 385.71 Proline--tRNA ligase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = proS PE = 3SV = 1 - [SYP_STRPS] B2INH5 83.95 31 299 370.13 SPFH domain/Band 7family OS = N- Streptococcus pneumoniae (strain terminally CGSP14) GN =SPCG_2124 PE = 4 SV = 1 - anchored [B2INH5_STRPS] (No CS) B2IQ40 64.1446 898 367.23 Phosphoenolpyruvate carboxylase Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = ppc PE = 3 SV = 1 -[B2IQ40_STRPS] B2IML8 76.35 25 406 358.33 Acetate kinase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = ackA PE = 3SV = 1 - [B2IML8_STRPS] B2IRS1 53.94 28 419 341.4 Cell division proteinFtsZ OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =ftsZ PE = 3 SV = 1 - [B2IRS1_STRPS] B2IMN0 96.59 22 352 339.61 Alcoholdehydrogenase, zinc- Intracellular containing OS =- Streptococcuspneumoniae (strain CGSP14) GN = adh PE = 3 SV = 1 - [B2IMNO_STRPS]B2ISJ7 51.09 13 137 337.74 30S ribosomal protein S12 OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = rpsL PE = 3 SV = 1 -[RS12_STRPS] B2IS41 63.29 16 207 337.71 50S ribosomal protein L4 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rplD PE = 3SV = 1 - [RL4_STRPS] B2ILW7 80.68 18 414 332.023-oxoacyl-[acyl-carrier-protein] Intracellular synthase 2 OS =-Streptococcus pneumoniae (strain CGSP14) GN = fabF PE = 3 SV = 1 -[B2ILW7_STRPS] B2IRM9 70.29 22 276 330.56 ABC transporter,substrate-binding Lipid protein OS =- Streptococcus anchored pneumoniae(strain CGSP14) GN = SPCG_0152 PE = 4 SV = 1 - [B2IRM9_STRPS] B2ISY947.99 21 498 319.21 Oligopeptide ABC transporter, Multi- permeaseprotein Amic OS = transmembrane Streptococcus pneumoniae (strain CGSP14)GN = amiC PE = 3 SV = 1 - [B2ISY9_STRPS] B2IQT6 62.75 22 459 313.52 NADHoxidase OS =- Streptococcus Intracellular pneumoniae (strain CGSP14) GN= nox PE = 4 SV = 1 - [B2IQT6_STRPS] B21PP2 47.02 32 721 312 Amino acidABC transporter, amino Multi- acid-binding protein/permeasetransmembrane protein OS =- Streptococcus pneumoniae (strain CGSP14) GN= SPCG_1062 PE = 3 SV = 1 - [B2IPP2_STRPS] B2IS47 63.5 13 137 300.82 50Sribosomal protein L16 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = rplP PE = 3 SV = 1 - [RL16_STRPS] B2IRC1 66.67 19339 299.62 UDP-glucose 4-epimerase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = galE PE = 4 SV = 1 - [B2IRC1_STRPS]B2IQW3 71.22 25 278 296.15 Amino acid ABC transporter, amino Lipidacid-binding protein OS = anchored Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1485 PE = 4 SV = 1 - [B2IQW3_STRPS] B2IRR1 67.83 19230 295.06 2,3-bisphosphoglycerate- Intracellular dependentphosphoglycerate mutase OS =- Streptococcus pneumoniae (strain CGSP14)GN = gpmA PE = 3 SV = 1 - [GPMA_STRPS] B2IS34 61.87 39 737 290.83Anaerobic ribonucleoside Intracellular triphosphate reductase OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0212 PE = 4 SV = 1 -[B2IS34_STRPS] B2IS52 87.78 18 180 288.42 50S ribosomal protein L5 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rplE PE = 3SV = 1 - [RL5_STRPS] B2IQU1 65.63 33 678 288.36 Glycine--tRNA ligasebeta subunit Intracellular OS =- Streptococcus pneumoniae (strainCGSP14) GN = glyS PE = 3 SV = 1 - [SYGB_STRPS] B2ILY5 60.36 21 555284.08 Uncharacterized protein OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0438 PE = 4 SV = 1 - [B2ILY5_STRPS]B2IS40 59.62 14 208 283.69 50S ribosomal protein L3 OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = rplC PE = 3 SV = 1 -[RL3_STRPS] B 21M73 32.44 18 521 281.46 Amino acid ABC transporter,amino Multi- acid-binding protein/permease transmembrane protein OS =-Streptococcus pneumoniae (strain CGSP14) GN = gInH PE = 3 SV = 1 -[B2IM73_STRPS] B2IR19 52.65 17 302 275.76 Amino acid ABC transporter,Lipid periplasmic amino acid-binding anchored protein, putative OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0108 PE = 4 SV = 1 -[B2IR19_STRPS] B2IRN0 57.75 22 284 275.2 Lipoprotein OS =- StreptococcusLipid pneumoniae (strain CGSP14) anchored GN = SPCG_0153 PE = 3 SV = 1 -[B2IRNO_STRPS] B2IM81 70.93 36 774 270.81 Formate acetyltransferase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = pfl PE = 4SV = 1 - [B2IM81_STRPS] B2IRN7 65.61 30 660 269.02 Threonine--tRNAligase OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =thrS PE = 3 SV = 1 - [B2IRN7_STRPS] B2IMN6 63.36 22 494 268.42 Threoninesynthase OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN= thrC PE = 4 SV = 1 - [B2IMN6_STRPS] B2IR98 69.68 20 376 261.66 SugarABC transporter, ATP-binding Intracellular protein OS =- Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1565 PE = 3 SV = 1 - [B2IR98_STRPS]B2IMG8 58.42 23 404 253.09 Aspartate aminotransferase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = aspC PE = 4 SV = 1 -[B2IMG8_STRPS] B2ILW6 65.43 13 243 252.823-ketoacyl-(Acyl-carrier-protein) Intracellular reductase OS =-Streptococcus pneumoniae (strain CGSP14) GN = fabG PE = 4 SV = 1 -[B2ILW6_STRPS] B2INF2 70.18 25 513 248.48 ABC transporter, ATP-bindingIntracellular protein OS =- Streptococcus pneumoniae (strain CGSP14) GN= SPCG_0719 PE = 4 SV = 1 - [B2INF2_STRPS] B2IRS2 78.12 20 457 247.37Cell division protein FtsA OS = Multi- Streptococcus pneumoniae (straintransmembrane CGSP14) GN = ftsZ PE = 3 SV = 1 - [B2IRS2_STRPS] B2IM4170.82 22 377 246.54 Uncharacterized protein OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = SPCG_1926 PE = 3 SV = 1 -[B2IM41_STRPS] B2IQ88 66.09 13 115 240.71 50S ribosomal protein L19 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rplS PE = 3SV = 1 - [RL19_STRPS] B2IQ10 56.1 22 344 240.32 Uncharacterized proteinOS = Multi- Streptococcus pneumoniae (strain transmembrane CGSP14) GN =SPCG_1352 PE = 4 SV = 1 - [B2IQ10_STRPS] B2IND2 63.73 18 386 236.96Branched-chain amino acid ABC Lipid transporter, amino acid-bindinganchored protein OS =- Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0699 PE = 4 SV = 1 - [B2IND2_STRPS] B2INP5 71.34 24 492 236.94Inosine-5′-monophosphate Intracellular dehydrogenase OS =- Streptococcuspneumoniae (strain CGSP14) GN = imdH PE = 3 SV = 1 - [B2INP5_STRPS]B2ISL2 67.26 18 339 235.53 Alcohol dehydrogenase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = adhP PE = 3 SV = 1 -[B2ISL2_STRPS] B2ISM4 59.23 9 130 228.51 30S ribosomal protein S9 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpsI PE = 3SV = 1 - [RS9_STRPS] B2IR71 40.26 23 914 227.01 Cation-transportingATPase, E1-E2 Multi- family OS =- Streptococcus pneumoniae transmembrane(strain CGSP14) GN = pacL PE = 3 SV = 1 - [B2IR71_STRPS] B2IPP1 85.77 17246 225.41 Amino acid ABC transporter, ATP- Intracellular bindingprotein OS =- Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1061 PE= 4 SV = 1 - [B2IPP1_STRPS] B2ISY7 58.31 18 355 221.22 Oligopeptide ABCtransporter, ATP- Intracellular binding protein AmiE OS = Streptococcuspneumoniae (strain CGSP14) GN = amiE PE = 3 SV = 1 - [B2ISY7_STRPS]B2IQL1 56.83 16 271 219.73 Amino acid ABC transporter, amino Lipidacid-binding protein OS = anchored Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1383 PE = 4 SV = 1 - [B2IQL1_STRPS] B2ISG3 77.23 19202 219.45 General stress protein 24, putative Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1781 PE = 4 SV = 1 -[B2ISG3_STRPS] B2IMA2 70.1 20 398 216.35 Phosphoglycerate kinase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = pgk PE = 3SV = 1 - [PGK_STRPS] B2IN71 49.03 25 620 212.01 Elongation factor Tufamily protein Intracellular OS =- Streptococcus pneumoniae (strainCGSP14) GN = SPCG_0638 PE = 4 SV = 1 - [B2IN71_STRPS] B2IPR1 75.6 16 332210.14 L-lactate dehydrogenase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = Idh PE = 3 SV = 1 - [B2IPR1_STRPS]B2IQK0 51.83 30 872 209.16 Alanine--tRNA ligase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = alaS PE = 3 SV = 1 -[SYA_STRPS] B2IS44 61.29 12 93 208.67 30S ribosomal protein S19 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpsS PE = 3SV = 1 - [RS19_STRPS] B2IMN9 48.47 17 425 208.24 Uncharacterized proteinOS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =SPCG_2034 PE = 4 SV = 1 - [B2IMN9_STRPS] B2IR17 72.27 16 339 208.13Ribose-phosphate Intracellular pyrophosphokinase OS = Streptococcuspneumoniae (strain CGSP14) GN = prsA PE = 3 SV = 1 - [B2IR17_STRPS]B2IQR1 62.69 25 520 208 GMP synthase [glutamine- Intracellularhydrolyzing] OS =- Streptococcus pneumoniae (strain CGSP14) GN = guaA PE= 3 SV = 1 - [GUAA_STRPS] B2INL3 50.25 23 810 202.66 ATP-dependent Clpprotease, ATP- Intracellular binding subunit OS =- Streptococcuspneumoniae (strain CGSP14) GN = SPCG_2162 PE = 3 SV = 1 - [B2INL3_STRPS]B2IN11 58.53 17 299 197.31 UTP--glucose-1-phosphate Secretoryuridylyltransferase OS = (released) Streptococcus pneumoniae (strain(with CS) CGSP14) GN = galU PE = 3 SV = 1 - [B2IN11_STRPS] B2IQU5 44.3817 480 197.17 Peptidoglycan N-acetylglucosamine N- deacetylase A OS =-Streptococcus terminally pneumoniae (strain CGSP14) anchored GN = pgdAPE = 4 SV = 1 - (with CS) [B2IQU5_STRPS] B2INE4 57.49 19 447 196.45DEAD-box ATP-dependent RNA Intracellular helicase CshB OS =-Streptococcus pneumoniae (strain CGSP14) GN = rheB PE = 3 SV = 1 -[B2INE4_STRPS] B2IMU6 65.97 15 238 196.17 Uncharacterized protein OS =Lipid Streptococcus pneumoniae (strain anchored CGSP14) GN = SPCG_0590PE = 4 SV = 1 - [B2IMU6_STRPS] B2INT7 54.92 27 752 195.47 ATP-dependentClp protease, ATP- Intracellular binding subunit ClpE OS = Streptococcuspneumoniae (strain CGSP14) GN = clpE PE = 3 SV = 1 - [B2INT7_STRPS]B2IM28 44.59 17 388 194.98 Protein RecA OS =- StreptococcusIntracellular pneumoniae (strain CGSP14) GN = recA PE = 3 SV = 1 -[RECA_STRPS] B2IQL4 79.78 18 267 192.86 Phosphate import ATP-bindingIntracellular protein PstB OS =- Streptococcus pneumoniae (strainCGSP14) GN = pstB PE = 3 SV = 1 - [B2IQL4_STRPS] B2ISL0 43.67 13 332190.43 PTS system, mannose-specific IIAB Intracellular components OS =-Streptococcus pneumoniae (strain CGSP14) GN = manL PE = 4 SV = 1 -[B2ISLO_STRPS] B2INW3 54.6 19 511 188.38 Sugar ABC transporter,ATP-binding Intracellular protein OS =- Streptococcus pneumoniae (strainCGSP14) GN = SPCG_0788 PE = 4 SV = 1 - [B2INW3_STRPS] B2INT0 40.9 22 648185.21 DNA gyrase subunit B OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = gyrB PE = 3 SV = 1 - [B2INTO_STRPS] B2IP17 57.18 13341 184.52 Iron-compound ABC transporter, Lipid iron compound-bindingprotein OS = anchored Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1009 PE = 4 SV = 1 - [B2IPI7_STRPS] B2IPT0 48.09 22 628 183.98Elongation factor 4 OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = lepA PE = 3 SV = 1 - [B2IPTO_STRPS] B2IN23 39.89 17 752181.97 Alpha-1,4 glucan phosphorylase Intracellular OS =- Streptococcuspneumoniae (strain CGSP14) GN = maIP PE = 3 SV = 1 - [B2IN23_STRPS]B2IQZ2 64.71 20 374 180.03 Ribosome-binding ATPase YchF OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = ychF PE = 3SV = 1 - [B2IQZ2_STRPS] B2IMU8 66.81 17 229 179.85 50S ribosomal proteinL1 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rplAPE = 3 SV = 1 - [RL1_STRPS] B2ISY6 51.06 14 331 178.98 Oligopeptide ABCtransporter, ATP- Intracellular binding protein AmiF OS = Streptococcuspneumoniae (strain CGSP14) GN = amiF PE = 3 SV = 1 - [B2ISY6_STRPS]B2IR87 54.15 18 410 170.5 ATP-dependent Clp protease ATP- Intracellularbinding subunit ClpX OS = Streptococcus pneumoniae (strain CGSP14) GN =clpX PE = 3 SV = 1 - [CLPX_STRPS] B2IR07 58.6 24 442 169.98Adenylosuccinate synthetase OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = purA PE = 3 SV = 1 - [B2IR07_STRPS] B2ILV9 61.23 20454 169.93 Aspartokinase OS =- Streptococcus Intracellular pneumoniae(strain CGSP14) GN = lysC PE = 3 SV = 1 - [B2ILV9_STRPS] B2IMZ6 42.12 25857 169.84 DNA mismatch repair protein MutS Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = hexA PE = 3 SV = 1 -[B2IMZ6_STRPS] B2IS57 84.76 17 164 169.34 30S ribosomal protein S5 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpsE PE = 3SV = 1 - [B2IS57_STRPS] B2ILR1 29.62 20 719 166.63 Penicillin-bindingprotein 1A OS = N- Streptococcus pneumoniae (strain terminally CGSP14)GN = pbp1A PE = 4 SV = 1 - anchored [B2ILR1_STRPS] (with CS) B2IQ7853.73 22 523 165.23 Signal recognition particle protein Intracellular OS=- Streptococcus pneumoniae (strain CGSP14) GN = ffh PE = 3 SV = 1 -[B2IQ78_STRPS] B2ISM3 78.38 17 148 163.82 50S ribosomal protein L13 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rplM PE = 3SV = 1 - [B2ISM3_STRPS] B2INJ2 70.02 21 427 163.61 DltD protein OS =-Streptococcus N- pneumoniae (strain CGSP14) terminally GN = dltD PE = 4SV = 1 - anchored [B2INJ2_STRPS] (with CS) B2ISJ1 48.84 19 602 159.21Glutamine--fructose-6-phosphate Intracellular aminotransferase[isomerizing] OS = Streptococcus pneumoniae (strain CGSP14) GN = glmS PE= 3 SV = 1 - [B2ISJ1_STRPS] B2IPM7 52.69 18 577 159.04Phosphoenolpyruvate-protein Intracellular phosphotransferase OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1121 PE = 3 SV = 1 -[B2IPM7_STRPS] B2IQ29 61.01 17 436 157.98 GTPase Obg OS =- StreptococcusIntracellular pneumoniae (strain CGSP14) GN = obg PE = 3 SV = 1 -[OBG_STRPS] B2IS66 74.8 9 127 157.55 30S ribosomal protein S11 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpsK PE = 3SV = 1 - [RS11_STRPS] B2IQ67 33.36 24 1058 157.35 Carbamoyl-phosphatesynthase Intracellular large chain OS =- Streptococcus pneumoniae(strain CGSP14) GN = carB PE = 3 SV = 1 - [CARB_STRPS] B2IQ25 62.29 16419 154.75 UDP-N-acetylglucosamine 1- Intracellularcarboxyvinyltransferase OS = Streptococcus pneumoniae (strain CGSP14) GN= murZ PE = 3 SV = 1 - [B2IQ25_STRPS] B2IQX4 82.93 14 164 154.18 ATPsynthase subunit b OS = N- Streptococcus pneumoniae (strain terminallyCGSP14) GN = atpF PE = 3 SV = 1 - anchored [ATPF_STRPS] (with CS) B2IM7269.51 10 246 154.11 Amino acid ABC transporter, ATP- Intracellularbinding protein OS =- Streptococcus pneumoniae (strain CGSP14) GN = glnQPE = 4 SV = 1 - [B2IM72_STRPS] B2INR2 35.79 19 679 153.3Methionine--tRNA ligase OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = metG PE = 3 SV = 1 - [B2INR2_STRPS] B2IS42 81.63 698 152.99 50S ribosomal protein L23 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = rplW PE = 3 SV = 1 - [RL23_STRPS] B2IPI044.02 18 418 152.53 Serine hydroxymethyltransferase Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = glyA PE = 3 SV = 1 -[GLYA_STRPS] B2IRG4 73.89 21 203 151.28 30S ribosomal protein S4 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpsD PE = 3SV = 1 - [RS4_STRPS] B2ILV7 59.29 21 452 150.33 Serine--tRNA ligase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = serS PE = 3SV = 1 - [B2ILV7_STRPS] B2INC8 60.65 13 216 146.88 Uracilphosphoribosyltransferase Intracellular OS =- Streptococcus pneumoniae(strain CGSP14) GN = upp PE = 3 SV = 1 - [B2INC8_STRPS] B2IQL3 70.63 16252 145.61 Phosphate import ATP-binding Intracellular protein PstB OS =-Streptococcus pneumoniae (strain CGSP14) GN = pstB PE = 3 SV = 1 -[B2IQL3_STRPS] B2IMZ5 31.91 19 564 144.21 ABC transporter, ATP- Multi-binding/permease protein OS = transmembrane Streptococcus pneumoniae(strain CGSP14) GN = SPCG_2042 PE = 4 SV = 1 - [B2IMZ5_STRPS] B2IS3959.8 8 102 143.18 30S ribosomal protein S10 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rpsJ PE = 3 SV = 1 -[RS10_STRPS] B2IP78 53.87 13 375 141.59 CarboxynorspermidineIntracellular decarboxylase OS =- Streptococcus pneumoniae (strainCGSP14) GN = nspC PE = 4 SV = 1 - [B2IP78_STRPS] B2IM60 54.81 13 416139.41 Diaminopimelate decarboxylase Intracellular OS =- Streptococcuspneumoniae (strain CGSP14) GN = lysA PE = 3 SV = 1 - [B2IM60_STRPS]B2IMH3 65.03 14 346 139.16 Catabolite control protein A OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = ccpA PE = 4SV = 1 - [B2IMH3_STRPS] B2IS68 57.03 8 128 139.07 50S ribosomal proteinL17 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =rplQ PE = 3 SV = 1 - [RL17_STRPS] B2INE0 34.95 13 329 136.77 Celldivision protein FtsX OS = Multi- Streptococcus pneumoniae (straintransmembrane CGSP14) GN = ftsX PE = 3 SV = 1 - [B2INEO_STRPS] B2ILV241.65 20 778 134.89 Endonuclease MutS2 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = mutS2 PE = 3 SV = 1 - [MUTS2_STRPS]B2IQU4 63.93 15 280 133.96 Oxidoreductase, aldo/keto Intracellularreductase family OS =- Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1466 PE = 4 SV = 1 - [B2IQU4_STRPS] B2IS55 71.35 14 178 129.78 50Sribosomal protein L6 OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = rplF PE = 3 SV = 1 - [RL6_STRPS] B2ISJ8 67.95 15 156 129.1630S ribosomal protein S7 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = rpsG PE = 3 SV = 1 - [RS7_STRPS] B2IRR6 24.81 8 266128.96 Cell division protein DivIVA OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1633 PE = 4 SV = 1 - [B2IRR6_STRPS]B2IS18 43.68 14 419 128.49 Zinc metalloprotease OS = Multi-Streptococcus pneumoniae (strain transmembrane CGSP14) GN = eep PE = 3SV = 1 - [B2IS18_STRPS] B2INF3 47.94 9 267 128.19 Peptidyl-prolylcis-trans isomerase Lipid OS =- Streptococcus pneumoniae (strainanchored CGSP14) GN = ppiA PE = 3 SV = 1 - [B2INF3_STRPS] B2IR96 96.5913 176 127.15 Adenine phosphoribosyltransferase Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = apt PE = 3 SV = 1 -[B2IR96_STRPS] B2IQX1 63.36 12 292 124.75 ATP synthase gamma chain OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = atpG PE = 3SV = 1 - [ATPG_STRPS] B2IQL7 40.41 12 292 124.49 Phosphate ABCtransporter, Lipid phosphate-binding protein, anchored putative OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1389 PE = 4 SV = 1 -[B2IQL7_STRPS] B2IS49 69.77 10 86 124.47 30S ribosomal protein S17 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpsQ PE = 3SV = 1 - [RS17_STRPS] B2IQV3 35.99 8 289 123.64 Glycerol uptakefacilitator protein, Multi- putative OS =- Streptococcus transmembranepneumoniae (strain CGSP14) GN = glpF PE = 3 SV = 1 - [B2IQV3_STRPS]B21M98 46.54 14 535 123.63 CTP synthase OS = Streptococcus Intracellularpneumoniae (strain CGSP14) GN = pyrG PE = 3 SV = 1 - [B2IM98_STRPS]B2IQ39 47.38 9 344 123.25 Uncharacterized protein OS = SecretoryStreptococcus pneumoniae (strain (released) CGSP14) GN = SPCG_1211 PE =4 SV = 1 - (with CS) [B2IQ39_STRPS] B2IP00 65.99 14 397 123.01 Serineprotease OS =- Streptococcus N- pneumoniae (strain CGSP14) terminally GN= sphtra PE = 4 SV = 1 - anchored [B2IP00_STRPS] (No CS) B2IN41 35.2 16429 122.13 Histidine--tRNA ligase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = hisS PE = 3 SV = 1 - [SYH_STRPS] B2IRW440.6 15 436 120.41 GTPase Der OS =- Streptococcus Intracellularpneumoniae (strain CGSP14) GN = der PE = 3 SV = 1 - [DER_STRPS] B2IR6835.97 13 531 119.86 Uncharacterized protein OS = Multi- Streptococcuspneumoniae (strain transmembrane CGSP14) GN = SPCG_1535 PE = 4 SV = 1 -[B2IR68_STRPS] B2IPR2 27.94 18 841 118.97 DNA gyrase subunit A OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = gyrA PE = 3SV = 1 - [B2IPR2_STRPS] B2IRA1 44.08 17 524 118.46 DEAD-boxATP-dependent RNA Intracellular helicase CshA OS =- Streptococcuspneumoniae (strain CGSP14) GN = cshA PE = 3 SV = 1 - [B2IRA1_STRPS]B2IM46 55.94 11 345 117.68 Uncharacterized protein OS = N- Streptococcuspneumoniae (strain terminally CGSP14) GN = SPCG_1931 PE = 4 SV = 1 -anchored [B2IM46_STRPS] (with CS) B2ILR7 48.44 16 481 117.486-phosphogluconate Intracellular dehydrogenase, decarboxylating OS =-Streptococcus pneumoniae (strain CGSP14) GN = gnd PE = 3 SV = 1 -[B2ILR7_STRPS] B2IR57 62.02 19 466 116.97 Cof familyprotein/peptidyl-prolyl Intracellular cis-trans isomerase, cyclophilintype OS =- Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1524 PE =4 SV = 1 - [B2IR57_STRPS] B2IND9 76.09 12 230 114.4 Cell division ABCtransporter, ATP- Intracellular binding protein FtsE OS = Streptococcuspneumoniae (strain CGSP14) GN = ftsE PE = 4 SV = 1 - [B2IND9_STRPS]B2IPU7 45 13 320 113.45 Ribonucleoside-diphosphate Intracellularreductase subunit beta OS = Streptococcus pneumoniae (strain CGSP14) GN= nrdF PE = 3 SV = 1 - [B2IPU7_STRPS] B2IMS9 43.58 20 553 112.79Ribonuclease J OS =- Streptococcus Intracellular pneumoniae (strainCGSP14) GN = rnj PE = 3 SV = 1 - [B2IMS9_STRPS] B2ISQ9 52.02 15 494111.9 UPF0371 protein SPCG_0344 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0344 PE = 3 SV = 1 - [Y344_STRPS]B2IPF4 53.38 14 311 110.86 Adhesion lipoprotein OS = Lipid Streptococcuspneumoniae (strain anchored CGSP14) GN = Imb PE = 3 SV = 1 -[B2IPF4_STRPS] B2IM45 50.82 12 427 110.79 UDP-N-acetylglucosamine 1-Intracellular carboxyvinyltransferase OS = Streptococcus pneumoniae(strain CGSP14) GN = murA PE = 3 SV = 1 - [B2IM45_STRPS] B2INN5 58.46 10272 109.88 Cell shape-determining protein N- MreC OS =- Streptococcuspneumoniae terminally (strain CGSP14) GN = mreC PE = 3 anchored SV = 1 -[B2INN5_STRPS] (with CS) B2IM15 46.84 10 158 109.73 Uncharacterizedprotein OS = Multi- Streptococcus pneumoniae (strain transmembraneCGSP14) GN = SPCG_1901 PE = 4 SV = 1 - [B2IM15_STRPS] B2IRP9 25.68 14740 109.17 GTP pyrophosphokinase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = reIA PE = 3 SV = 1 - [B2IRP9_STRPS]B2INE1 34.71 11 726 108.73 PTS system, IIABC components OS = Multi-Streptococcus pneumoniae (strain transmembrane CGSP14) GN = ptsG PE = 4SV = 1 - [B2INE1_STRPS] B2IST7 48.44 12 417 108.26 Capsularpolysaccharide Intracellular biosynthesis protein, putative OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1812 PE = 3 SV = 1 -[B2IST7_STRPS] B2IP76 71.33 14 286 106.74 Polyamineaminopropyltransferase Intracellular OS =- Streptococcus pneumoniae(strain CGSP14) GN = speE PE = 3 SV = 1 - [B2IP76_STRPS] B2IS23 34.49 22954 105.34 UvrABC system protein A OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = uvrA PE = 3 SV = 1 - [B2IS23_STRPS]B2IS53 52.81 7 89 105.15 30S ribosomal protein S14 OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = rpsN PE = 3 SV = 1 -[RS14_STRPS] B2IQN9 49.79 20 486 105.03 Nicotinate Intracellularphosphoribosyltransferase OS = Streptococcus pneumoniae (strain CGSP14)GN = SPCG_1408 PE = 3 SV = 1 - [B2IQN9_STRPS] B2ISZ6 49.88 13 419 104.34Sugar ABC transporter, sugar- Lipid binding protein OS =- Streptococcusanchored pneumoniae (strain CGSP14) GN = SPCG_1871 PE = 4 SV = 1 -[B2ISZ6_STRPS] B2IMA9 47.02 17 487 101.63 Type Irestriction-modification Intracellular system, M subunit OS =Streptococcus pneumoniae (strain CGSP14) GN = hsdM PE = 4 SV = 1 -[B2IMA9_STRPS] B2IQH2 72.19 13 187 100.73 50S ribosomal protein L10 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rplJ PE = 3SV = 1 - [B2IQH2_STRPS] B2IR61 39.82 14 447 100.57 Asparagine--tRNAligase OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =asnS PE = 3 SV = 1 - [B2IR61_STRPS] B2IM30 50 15 338 100.22Transcriptional regulator, putative Secretory OS =- Streptococcuspneumoniae (strain (released) CGSP14) GN = lytR PE = 4 SV = 1 - (withCS) [B2IM30_STRPS] B2ISV9 32.29 10 511 99.16 Choline transporter OS =Multi- Streptococcus pneumoniae (strain transmembrane CGSP14) GN =SPCG_1834 PE = 3 SV = 1 - [B2ISV9_STRPS] B2IS51 78.22 12 101 98.15 50Sribosomal protein L24 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = rplX PE = 3 SV = 1 - [RL24_STRPS] B2IRX0 76.67 11240 97.84 ABC transporter, ATP-binding Intracellular protein OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1687 PE = 4 SV = 1 -[B2IRX0_STRPS] B2ISX8 46.73 11 336 97.53 Non-canonical purine NTPIntracellular pyrophosphatase OS =- Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1853 PE = 3 SV = 1 - [B2ISX8_STRPS] B2IN24 45.35 14505 97.46 4-alpha-glucanotransferase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = maIM PE = 3 SV = 1 - [B2IN24_STRPS]B2IM61 54.55 12 275 97.4 Purine operon repressor OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = purR PE = 4 SV = 1 -[B2IM61_STRPS] B2ILX0 55.38 16 455 97.1 Acetyl-CoA carboxylase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = accC PE = 4SV = 1 - [B2ILXO_STRPS] B2IQ19 35.91 15 763 96.38 DNA helicase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = pcrA PE = 3SV = 1 - [B2IQ19_STRPS] B2IPZ6 69.23 10 91 95.95 DNA-binding protein HUOS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = hlpA PE= 3 SV = 1 - [B2IPZ6_STRPS] B2INE5 31.57 12 396 95.45S-adenosylmethionine synthase Intracellular OS =- Streptococcuspneumoniae (strain CGSP14) GN = metK PE = 3 SV = 1 - [METK_STRPS] B2IS8228.53 15 659 94.22 Serine/threonine protein kinase Intracellular/ OS =-Streptococcus pneumoniae (strain TMH start CGSP14) GN = pkn2 PE = 3 SV =1 - AFTER 60 [B2IS82_STRPS] B2IS65 58.68 10 121 93.19 30S ribosomalprotein S13 OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = rpsM PE = 3 SV = 1 - [RS13_STRPS] B21MI4 35.02 18 731 92.99Penicillin-binding protein 2A OS = N- Streptococcus pneumoniae (strainterminally CGSP14) GN = pbp2A PE = 4 SV = 1 - anchored [B2IMI4_STRPS](with CS) B2IRK4 37.05 17 637 92.87 tRNA uridine 5- Intracellularcarboxymethylaminomethyl modification enzyme MnmG OS = Streptococcuspneumoniae (strain CGSP14) GN = mnmG PE = 3 SV = 1 - [B2IRK4_STRPS]B2ISW0 69.01 11 242 92.71 Choline transporter OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = SPCG_1835 PE = 4 SV = 1 -[B2ISWO_STRPS] B2INP6 39.59 10 341 92.59 Tryptophan--tRNA ligase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = trpS PE = 3SV = 1 - [B2INP6_STRPS] B2INC5 39.5 11 281 92.15 Uncharacterized proteinOS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0692 PE = 4 SV = 1 - [B2INC5_STRPS] B2IP66 36.64 17 715 91.51Transcriptional regulator, putative Intracellular OS =- Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0885 PE = 4 SV = 1 - [B2IP66_STRPS]B2ISY1 46.58 14 541 91.27 Dextran glucosidase DexS, putativeIntracellular OS =- Streptococcus pneumoniae (strain CGSP14) GN = dexSPE = 4 SV = 1 - [B2ISY1_STRPS] B2ISK7 38.51 11 444 91.26 AminopeptidaseC OS =- Streptococcus Intracellular pneumoniae (strain CGSP14) GN = pepCPE = 4 SV = 1 - [B2ISK7_STRPS] B2INL6 30.75 8 335 91.2 ABC transporter,substrate-binding Lipid protein, putative OS =- Streptococcus anchoredpneumoniae (strain CGSP14) GN = SPCG_2165 PE = 4 SV = 1 - [B2INL6_STRPS]B2IQY9 25.83 11 453 90.37 Chromosomal replication initiatorIntracellular protein DnaA OS =- Streptococcus pneumoniae (strainCGSP14) GN = dnaA PE = 3 SV = 1 - [DNAA_STRPS] B2IRS8 22.34 13 685 90.12Penicillin-binding protein 2B OS = N- Streptococcus pneumoniae (strainterminally CGSP14) GN = pbp2B PE = 4 SV = 1 - anchored [B2IRS8_STRPS](with CS) B2IS56 53.44 8 131 88.46 50S ribosomal protein L18 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rplR PE = 3SV = 1 - [B2IS56_STRPS] B2IQ12 40.13 9 319 88.41 Ribose-phosphateIntracellular pyrophosphokinase OS = Streptococcus pneumoniae (strainCGSP14) GN = prs PE = 3 SV = 1 - [B2IQ12_STRPS] B2IS67 52.73 9 311 88.17DNA-directed RNA polymerase Intracellular subunit alpha OS =-Streptococcus pneumoniae (strain CGSP14) GN = rpoA PE = 3 SV = 1 -[B2IS67_STRPS] B2IS59 64.38 9 146 88.05 50S ribosomal protein L15 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rplO PE = 3SV = 1 - [RL15_STRPS] B2IQ61 41.52 9 289 87.7 Choline kinase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = pck PE = 4SV = 1 - [B2IQ61_STRPS] B2ISY2 24.4 10 705 87.27 Trehalose PTS system,IIABC Multi- components OS =- Streptococcus transmembrane pneumoniaestrain CGSP14) GN = treP PE = 4 SV = 1 - [B2ISY2_STRPS] B2IMQ3 25.97 12801 86.8 Phenylalanine--tRNA ligase beta Intracellular subunit OS =-Streptococcus pneumoniae (strain CGSP14) GN = pheT PE = 3 SV = 1 -[B2IMQ3_STRPS] B2IQ02 60.82 9 97 86.49 50S ribosomal protein L27 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpmA PE = 3SV = 1 - [RL27_STRPS] B2IN58 40.3 11 335 86.4 Uncharacterized protein OS= Multi- Streptococcus pneumoniae (strain transmembrane CGSP14) GN =SPCG_2101 PE = 4 SV = 1 - [B2IN58_STRPS] B2IQW4 62.2 10 209 84.4 Aminoacid ABC transporter, ATP- Intracellular binding protein OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1486 PE = 4 SV = 1 -[B2IQW4_STRPS] B2IP74 35.23 13 491 84.24 Lysine decarboxylase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = cad PE = 4SV = 1 - [B2IP74_STRPS] B2IS70 42.49 11 433 83.11 UPF0210 proteinSPCG_0246 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN= SPCG_0246 PE = 3 SV = 1 - [B2IS70_STRPS] B2IQ66 64.29 13 280 83.05LicD2 protein OS =- Streptococcus Intracellular pneumoniae (strainCGSP14) GN = licD2 PE = 4 SV = 1 - [B2IQ66_STRPS] B2IRK5 34.17 12 55981.76 Ribonuclease J OS =- Streptococcus Intracellular pneumoniae(strain CGSP14) GN = rnj PE = 3 SV = 1 - [B2IRK5_STRPS] B2IR84 68.24 12296 81.42 Nucleotide-binding protein Intracellular SPCG_1551 OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1551 PE = 3 SV = 1 -[Y1551_STRPS] B2IMB5 34.93 15 607 81.24 Chaperone protein Dnak OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = dnaK PE = 3SV = 1 - [DNAK_STRPS] B2IQ62 58.09 12 346 80.31 Alcohol dehydrogenase,zinc- Intracellular containing OS =- Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1234 PE = 4 SV = 1 - [B2IQ62_STRPS] B2IMB9 37.86 11243 79.94 ABC transporter, ATP-binding Intracellular protein OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0494 PE = 4 SV = 1 -[B2IMB9_STRPS] B2IRD4 56.18 5 89 79.7 30S ribosomal protein S15 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpsO PE = 3SV = 1 - [RS15_STRPS] B2IN68 50.79 8 126 79.14 Uncharacterized proteinOS = N- Streptococcus pneumoniae (strain terminally CGSP14) GN =SPCG_0635 PE = 4 SV = 1 - anchored [B2IN68_STRPS] (with CS) BEIQN0 44.093 127 78.99 Uncharacterized protein OS = N- Streptococcus pneumoniae(strain terminally CGSP14) GN = SPCG_1399 PE = 4 SV = 1 anchored[B2IQNO_STRPS] (with CS) B2ILY0 36.58 14 514 78.96 Peptide chain releasefactor 3 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN= prfC PE = 3 SV = 1 - [RF3_STRPS] B2IS54 57.58 8 132 78.42 30Sribosomal protein S8 OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = rpsH PE = 3 SV = 1 - [RS8_STRPS] B2IMB6 35.19 13 378 78.27Chaperone protein DnaJ OS- Intracellular Streptococcus pneumoniae(strain CGSP14) GN = dnaJ PE = 3 SV = 1 - [B2IMB6_STRPS] B2IQS5 42.11 12247 78.16 Amino acid ABC transporter, ATP- Intracellular binding proteinOS =- Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1447 PE = 4 SV= 1 - [B2IQS5_STRPS] B2IRH0 33.01 12 512 76.84 ABC transporter,substrate-binding Lipid protein OS =- Streptococcus anchored pneumoniae(strain CGSP14) GN = SPCG_0089 PE = 4 SV = 1 - [B2IRHO_STRPS] B21PW950.88 11 283 76.72 Ribosome biogenesis GTPase A OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = SPCG_1141 PE = 3 SV = 1 -[B2IPW9_STRPS] B2IN18 48.09 11 418 76.44 Tyrosine--tRNA ligase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = tyrS PE = 3SV = 1 - [SYY_STRPS] B2IQV9 33.39 13 572 75.14 Phosphoglucomutase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = pgm PE = 3SV = 1 - [B2IQV9_STRPS] B2IP77 38.9 9 419 74.94 Uncharacterized proteinOS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0896 PE = 4 SV = 1 - [B2IP77_STRPS] B2IQ63 53.19 8 235 74.222-C-methyl-D-erythritol 4- Intracellular phosphate cytidylyltransferaseOS = Streptococcus pneumoniae (strain CGSP14) GN = ispD PE = 3 SV = 1 -[ISPD_STRPS] B2IRQ4 65.34 11 251 74.2 Manganese ABC transporter, ATP-Intracellular binding protein OS =- Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1621 PE = 4 SV = 1 - [B2IRQ4_STRPS] B2IQX3 62.36 8 17873.32 ATP synthase subunit delta OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = atpH PE = 3 SV = 1 - [ATPD_STRPS] B2INX427.48 12 826 73.21 DNA topoisomerase 4 subunit A Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = parC PE = 3 SV = 1 -[B2INX4_STRPS] B2IN76 40.67 12 450 72.88 UDP-N-acetylmuramoylalanine--D-Intracellular glutamate ligase OS =- Streptococcus pneumoniae (strainCGSP14) GN = murD PE = 3 SV = 1 - [MURD_STRPS] B2IPC3 41.47 10 299 72.75GTPase Era OS =- Streptococcus Intracellular pneumoniae (strain CGSP14)GN = era PE = 3 SV = 1 - [ERA_STRPS] B2IRX7 40.79 13 353 72.1 Methionineimport ATP-binding Intracellular protein MetN OS =- Streptococcuspneumoniae (strain CGSP14) GN = metN PE = 3 SV = 1 - [B2IRX7_STRPS]B2INU1 50.65 9 306 70.6 Bifunctional protein FoID OS = LipidStreptococcus pneumoniae (strain anchored CGSP14) GN = foID PE = 3 SV =1 - [B2INU1_STRPS] B2ILW4 36.11 6 324 69.62 Enoyl-(Acyl-carrier-protein)Intracellular reductase OS =- Streptococcus pneumoniae (strain CGSP14)GN = fabK PE = 4 SV = 1 - [B2ILW4_STRPS] B2IS45 40.35 3 114 69.56 50Sribosomal protein L22 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = rplV PE = 3 SV = 1 - [RL22_STRPS] B2IPZ1 30.21 13652 67.59 DNA ligase OS =- Streptococcus Intracellular pneumoniae(strain CGSP14) GN = ligA PE = 3 SV = 1 - [DNLJ_STRPS] B2IMZ3 34.66 12603 66.96 ABC transporter, ATP- Multi- binding/permease protein OS =transmembrane Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2040 PE= 4 SV = 1 - [B2IMZ3_STRPS] B2IS63 54.72 6 212 66.76 Adenylate kinase OS=- Streptococcus Intracellular pneumoniae (strain CGSP14) GN = adk PE =3 SV = 1 - [KAD_STRPS] B2INV5 53.01 6 83 65.46 30S ribosomal protein S20OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpsT PE= 3 SV = 1 - [B2INV5_STRPS] B2IRK2 38.07 10 373 65.45 tRNA-specific2-thiouridylase Intracellular MnmA OS =- Streptococcus pneumoniae(strain CGSP14) GN = mnmA PE = 3 SV = 2 - [MNMA_STRPS] B2IRB7 27.22 2158 65.21 Uncharacterized protein OS = N- Streptococcus pneumoniae(strain terminally CGSP14) GN = SPCG_1584 PE = 4 SV = 1 - anchored[B2IRB7_STRPS] (with CS) B2INE6 56.27 10 311 64.68 Dihydroorotatedehydrogenase OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = pyrD PE = 3 SV = 1 - [B2INE6_STRPS] B2IMB0 27.41 15 77764.13 Type I restriction-modification Intracellular system, R subunit OS=- Streptococcus pneumoniae (strain CGSP14) GN = hsdR PE = 4 SV = 1 -[B2IMBO_STRPS] B2ISQ4 20.27 11 750 63.95 Penicillin-binding protein 2XOS = N- Streptococcus pneumoniae (strain terminally CGSP14) GN = pbp2XPE = 4 SV = 1 - anchored [B2ISQ4_STRPS] (with CS) B2IPQ3 27.88 12 55663.73 Formate--tetrahydrofolate ligase Intracellular OS =- Streptococcuspneumoniae (strain CGSP14) GN = fhs PE = 3 SV = 1 - [FTHS_STRPS] B2IMU741.84 5 141 63.38 50S ribosomal protein L11 OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = rplK PE = 3 SV = 1 -[RL11_STRPS] B2ISY0 38 4 100 62.82 UPF0154 protein SPCG_1855 OS = N-Streptococcus pneumoniae (strain terminally CGSP14) GN = SPCG_1855 PE =3 SV = 1 - anchored [B2ISYO_STRPS] (with CS) B2IQP4 29.49 9 434 62.33Peptidase, U32 family OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1416 PE = 4 SV = 1 - [B2IQP4_STRPS] B2IR7957.14 11 259 62.31 Uncharacterized protein OS = N- Streptococcuspneumoniae (strain terminally CGSP14) GN = SPCG_1546 PE = 4 SV = 1 -anchored [B2IR79_STRPS] (with CS) B2ILW2 21.6 8 324 62.193-oxoacyl-[acyl-carrier-protein] Intracellular synthase 3 OS =-Streptococcus pneumoniae (strain CGSP14) GN = fabH PE = 3 SV = 1 -[FABH_STRPS] B2IPZ9 43.28 9 305 61.07 Riboflavin biosynthesis protein OS= Intracellular Streptococcus pneumoniae (strain CGSP14) GN = mreA PE =3 SV = 1 - [B2IPZ9_STRPS] B2IN32 29.98 13 587 60.72 Aspartate--tRNAligase OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =aspS PE = 3 SV = 1 - [SYD_STRPS] B2IRR9 42.46 7 179 60.72 Cell divisionprotein SepF OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = sepF PE = 3 SV = 1 - [SEPF_STRPS] B2IN19 28.12 13 690 60.07Cation-transporting ATPase, El-E2 Intracellular family OS =-Streptococcus pneumoniae (strain CGSP14) GN = ctpC PE = 3 SV = 1 -[B2IN19_STRPS] B2IQY8 36.71 9 444 59.95 UDP-N-acetylmuramate--L-alanineIntracellular ligase OS =- Streptococcus pneumoniae (strain CGSP14) GN =murC PE = 3 SV = 1 - [MURC_STRPS] B2IS26 33.86 8 189 59.81Uncharacterized protein OS = Lipid Streptococcus pneumoniae (strainanchored CGSP14) GN = SPCG_0204 PE = 4 SV = 1 [B2IS26_STRPS] B2INY745.25 8 263 59.27 ABC transporter, ATP-binding Intracellular protein OS=- Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0812 PE = 4 SV =1 - [B2INY7_STRPS] B2ILR6 14.66 6 464 58.78 Mid-cell-anchored protein ZOS = N- Streptococcus pneumoniae (strain terminally CGSP14) GN = mapZ PE= 3 SV = 1 - anchored [B2ILR6_STRPS] (with CS) B2ILX8 36.68 10 488 58.59Glutamyl-tRNA(Gln) Intracellular amidotransferase subunit A OS =Streptococcus pneumoniae (strain CGSP14) GN = gatA PE = 3 SV = 1 -[GATA_STRPS] B2IMN3 35.26 7 380 58.5 Queuine tRNA-ribosyltransferaseIntracellular OS =- Streptococcus pneumoniae (strain CGSP14) GN = tgt PE= 3 SV = 1 - [TGT_STRPS] B2IN17 21.09 12 825 58.4 Penicillin-bindingprotein 1B OS = Intracellular/ Streptococcus pneumoniae (strain TMHstart CGSP14) GN = pbp1B PE = 4 SV = 1 - AFTER 60 [B2IN17_STRPS] B2IR2745.45 8 330 57.61 Phosphate acyltransferase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = plsX PE = 3 SV = 1 -[PLSX_STRPS] B2INF7 73.21 6 112 56.77 ATP cone domain-containingIntracellular protein OS =- Streptococcus pneumoniae (strain CGSP14) GN= SPCG_0728 PE = 4 SV = 1 - [B2INF7_STRPS] B2IMZ8 20.78 9 563 56.13Arginine--tRNA ligase OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = argS PE = 3 SV = 1 - [SYR_STRPS] B2ILU5 35.6 10 42755.96 Trigger factor OS =- Streptococcus Intracellular pneumoniae(strain CGSP14) GN = tig PE = 3 SV = 1 - [TIG_STRPS] B2INK8 56.55 9 33655.86 tRNA-dihydrouridine synthase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = SPCG_2157 PE = 3 SV = 1 [B2INK8_STRPS]B2IM90 37.14 11 560 55.51 ABC transporter, ATP-binding Intracellularprotein OS =- Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0465 PE= 4 SV = 1 - [B2IM90_STRPS] B2IQA4 37.72 12 448 54.94 Glutamatedehydrogenase OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = gdhA PE = 3 SV = 1 - [B2IQA4_STRPS] B2IPP0 30.1 8 495 54.8Glucose-6-phosphate 1- Intracellular dehydrogenase OS =- Streptococcuspneumoniae (strain CGSP14) GN = zwf PE = 3 SV = 1 - [B2IPPO_STRPS]B2IPZ5 24.33 13 633 54.29 ABC transporter, ATP-binding Intracellularprotein OS =- Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1167 PE= 4 SV = 1 - [B2IPZ5_STRPS] B2IS10 23.73 8 649 54.27 DNA mismatch repairprotein MutL Intracellular OS =- Streptococcus pneumoniae (strainCGSP14) GN = mutL PE = 3 SV = 1 - [MUTL_STRPS] B2IN96 47.62 10 378 54.14Lactate oxidase OS =- Streptococcus Intracellular pneumoniae (strainCGSP14) GN = IctO PE = 4 SV = 1 - [B2IN96_STRPS] B2IND6 51.69 6 23653.89 Branched-chain amino acid ABC Intracellular transporter,ATP-binding protein OS =- Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0703 PE = 4 SV = 1 - [B2IND6_STRPS] B2IS98 58.85 9 209 53.7Probable nicotinate-nucleotide Intracellular adenylyltransferase OS =Streptococcus pneumoniae (strain CGSP14) GN = nadD PE = 3 SV = 1 -[B2IS98_STRPS] B2IPN8 43.01 8 272 53.21 Cof family protein OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = SPCG_1058 PE= 4 SV = 1 - [B2IPN8_STRPS] B2INN8 51.27 10 275 52.83 Energy-couplingfactor transporter N- ATP-binding protein EcfA OS = terminallyStreptococcus pneumoniae (strain anchored CGSP14) GN = ecfA PE = 3 SV =1 - (with CS) [B2INN8_STRPS] B2ISR0 23.55 8 535 52.43 Glucan1,6-alpha-glucosidase OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = dexB PE = 4 SV = 1 - [B2ISRO_STRPS] B2IRF6 52.19 8228 52.22 Potassium uptake protein, Trk N- family OS =- Streptococcuspneumoniae terminally (strain CGSP14) GN = trkA PE = 4 SV = 1 - anchored[B2IRF6_STRPS] (with CS) B2IM43 17.16 7 443 51.63 CBS domain protein OS= Multi- Streptococcus pneumoniae (strain transmembrane CGSP14) GN =SPCG_1928 PE = 4 SV = 1 - [B2IM43_STRPS] B2ILX2 40 6 255 51.58Acetyl-coenzyme A carboxylase Intracellular carboxyl transferase subunitalpha OS =- Streptococcus pneumoniae (strain CGSP14) GN = accA PE = 3 SV= 1 - [ACCA_STRPS] B2ISA0 34.78 10 368 51.32 GTP-binding protein OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1723 PE= 4 SV = 1 - [B2ISAO_STRPS] B2IPB9 23.19 10 677 50.7Endo-beta-N-acetylglucosaminidase OS =- Secretory Streptococcuspneumoniae (strain (released) CGSP14) GN = lytB PE = 4 SV = 1 - (withCS) [B2IPB9_STRPS] B2IQW9 61.87 6 139 50.69 ATP synthase epsilon chainOS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = atpC PE= 3 SV = 1 - [ATPE_STRPS] B2IPN9 24.94 7 429 50.34 Signal recognitionparticle receptor Intracellular FtsY OS =- Streptococcus pneumoniae(strain CGSP14) GN = ftsY PE = 3 SV = 1 - [B2IPN9_STRPS] B2IRI7 17.95 9713 49.57 Uncharacterized protein OS = Multi- Streptococcus pneumoniae(strain transmembrane CGSP14) GN = SPCG_0106 PE = 4 SV = 1 -[B2IRI7_STRPS] B2IQH3 24.39 7 488 49.39 Chlorohydrolase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = trzA PE = 4SV = 1 - [B2IQH3_STRPS] B2IRD2 21.59 7 264 48.68 Acyltransferase familyprotein OS = Multi- Streptococcus pneumoniae (strain transmembraneCGSP14) GN = SPCG_1599 PE = 4 SV = 1 - [B2IRD2_STRPS] B2IR63 31.14 7 39548.62 Aspartate aminotransferase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = aspB PE = 3 SV = 1 - [B2IR63_STRPS]B2INK4 23.36 10 608 48.46 Uncharacterized protein OS = N- Streptococcuspneumoniae (strain terminally CGSP14) GN = SPCG_2153 PE = 4 SV = 1 -anchored [B2INK4_STRPS] (with CS) B2IN18 28.14 7 501 48.06 Zinc ABCtransporter, zinc-binding Lipid lipoprotein OS =- Streptococcus anchoredpneumoniae (strain CGSP14) GN = SPCG_2137 PE = 3 SV = 1 - [B2IN18_STRPS]B2IPU4 40.71 7 253 47.82 Lactose phosphotransferase system Intracellularrepressor OS =- Streptococcus pneumoniae (strain CGSP14) GN = lacR PE =4 SV = 1 - [B2IPU4_STRPS] B2INN7 41.94 8 279 47.56 Energy-couplingfactor transporter N- ATP-binding protein EcfA OS = terminallyStreptococcus pneumoniae (strain anchored CGSP14) GN = ecfA PE = 3 SV =1 - (with CS) [B2INN7_STRPS] B2IS50 52.46 7 122 47.46 50S ribosomalprotein L14 OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = rplN PE = 3 SV = 1 - [RL14_STRPS] B2IRI6 32.63 3 95 47.46Bacteriocin, putative OS = N- Streptococcus pneumoniae (strainterminally CGSP14) GN = SPCG_0105 PE = 4 SV = 1 - anchored[B2IRI6_STRPS] (with CS) B2IPJ1 50 9 264 47.22 Iron-compound ABCtransporter, Intracellular ATP-binding protein OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1013 PE = 4 SV = 1 - [B2IPJ1_STRPS]B2IPR3 40.89 7 247 47.21 Uncharacterized protein OS = N- Streptococcuspneumoniae (strain terminally CGSP14) GN = SPCG_1083 PE = 4 SV = 1 -anchored [B2IPR3_STRPS] (with CS) B2IR24 28.28 9 389 47.15Uncharacterized protein OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = SPCG_0036 PE = 3 SV = 1 - [B2IR24_STRPS] B2ISX760.12 8 173 47.03 Phosphoesterase OS =- Streptococcus Intracellularpneumoniae (strain CGSP14) GN = SPCG_1852 PE = 3 SV = 1 - [B2ISX7_STRPS]B2IM86 29.62 8 449 46.75 Potassium uptake protein, Trk N- family OS =-Streptococcus pneumoniae terminally (strain CGSP14) GN = trkA PE = 4 SV= 1 - anchored [B2IM86_STRPS] (with CS) B2IPT8 33.13 9 326 46.7 Tagatose1,6-diphosphate aldolase Intracellular OS =- Streptococcus pneumoniae(strain CGSP14) GN = lacD PE = 3 SV = 1 - [B2IPT8_STRPS] B2ILZ5 39.81 10540 46.59 60 kDa chaperonin OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = groL PE = 3 SV = 1 - [CH60_STRPS] B2ILX7 34.58 8480 46.43 Aspartyl/glutamyl-tRNA(Asn/Gln) Intracellular amidotransferasesubunit B OS = Streptococcus pneumoniae (strain CGSP14) GN = gatB PE = 3SV = 1 - [GATB_STRPS] B2IMQ9 16.96 8 737 45.59 PolyribonucleotideIntracellular nucleotidyltransferase OS = Streptococcus pneumoniae(strain CGSP14) GN = pnp PE = 3 SV = 2 - [PNP_STRPS] B2IQN3 45.45 5 6644.91 30S ribosomal protein S21 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = rpsU PE = 3 SV = 1 - [B2IQN3_STRPS]B2IR90 38.37 2 172 44.68 Non-heme iron-containing ferritin IntracellularOS =- Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1557 PE = 3 SV= 1 - [B2IR90_STRPS] B2IMS2 42.66 7 293 44.59 Fructose-bisphosphatealdolase Intracellular OS =- Streptococcus pneumoniae (strain CGSP14) GN= fba PE = 3 SV = 1 - [B2IMS2_STRPS] B2IRX2 10.53 5 399 44.06Uncharacterized protein OS = Multi- Streptococcus pneumoniae (straintransmembrane CGSP14) GN = SPCG_1689 PE = 4 SV = 1 - [B2IRX2_STRPS]B2ISH7 20.98 10 815 43.34 Formate acetyltransferase, putativeIntracellular OS =- Streptococcus pneumoniae (strain CGSP14) GN = pfIFPE = 4 SV = 1 - [B2ISH7_STRPS] B2ISG1 26.84 7 190 43.06 Uncharacterizedprotein OS = Multi- Streptococcus pneumoniae (strain transmembraneCGSP14) GN = SPCG_1779 PE = 4 SV = 1 - [B2ISG1_STRPS] B21MY2 73.21 7 5642.86 Uncharacterized protein OS = Secretory Streptococcus pneumoniae(strain (released) CGSP14) GN = SPCG_0626 PE = 4 SV = 1 - (with CS)[B2IMY2_STRPS] B2IM36 15.5 8 671 42.81 Uncharacterized protein OS =Multi- Streptococcus pneumoniae (strain transmembrane CGSP14) GN =SPCG_1921 PE = 4 SV = 1 [B2IM36_STRPS] B2IM12 26.96 8 471 42.77Pneumolysin OS =- Streptococcus Intracellular pneumoniae (strain CGSP14)GN = ply PE = 4 SV = 1 - [B2IM12_STRPS] B2INM4 34.62 6 182 42.71Ribosomal subunit interface protein Intracellular OS =- Streptococcuspneumoniae (strain CGSP14) GN = SPCG_2173 PE = 4 SV = 1 [B2INM4_STRPS]B2IPW4 27.34 9 567 42.61 Dihydrolipoyl dehydrogenase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = acoL PE = 4 SV = 1 -[B2IPW4_STRPS] B2IQZ9 38.89 6 180 42.56 Hypoxanthine-guanineIntracellular phosphoribosyltransferase OS = Streptococcus pneumoniae(strain CGSP14) GN = hgt PE = 4 SV = 1 - [B2IQZ9_STRPS] B2IS21 19.23 6234 42.44 Uncharacterized protein OS = Multi- Streptococcus pneumoniae(strain transmembrane CGSP14) GN = SPCG_0199 PE = 4 SV = 1 -[B2IS21_STRPS] B2IPB4 18.18 3 66 42.39 50S ribosomal protein L35 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpml PE = 3SV = 1 - [RL35_STRPS] B2IM29 23.68 5 418 42.24 Putativecompetence-damage Intracellular inducible protein OS =- Streptococcuspneumoniae (strain CGSP14) GN = cinA PE = 3 SV = 1 - [CINA_STRPS] B2IR8666.5 11 197 41.89 Probable GTP-binding protein EngB Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = engB PE = 3 SV = 1 -[ENGB_STRPS] B2IPS2 46.7 4 212 41.29 Uridine kinase OS =- StreptococcusIntracellular pneumoniae (strain CGSP14) GN = udk PE = 3 SV = 1 -[URK_STRPS] B2INP0 22.46 5 276 41.12 Uncharacterized protein OS =Intracellular/ Streptococcus pneumoniae (strain TMH start CGSP14) GN =SPCG_2189 PE = 4 SV = 1 - AFTER 60 [B2INP0_STRPS] B2ILX1 42.71 7 28841.03 Acetyl-coenzyme A carboxylase Intracellular carboxyl transferasesubunit beta OS =- Streptococcus pneumoniae (strain CGSP14) GN = accD PE= 3 SV = 1 - [B2ILX1_STRPS] B2IMY9 31.48 10 486 40.7 Glutamate--tRNAligase OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =gltX PE = 3 SV = 1 - [SYE_STRPS] B2ISL4 51.11 9 270 40.64 Cof familyprotein OS =- Streptococcus Intracellular pneumoniae (strain CGSP14) GN= SPCG_0299 PE = 4 SV = 1 - [B2ISL4_STRPS] B2IQH7 42.21 5 289 40.55Homoserine kinase OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = thrB PE = 3 SV = 1 - [KHSE_STRPS] B2IS89 36.54 7 208 40.54Guanylate kinase OS =- Streptococcus Intracellular pneumoniae (strainCGSP14) GN = gmk PE = 3 SV = 1 - [B2IS89_STRPS] B2IRQ3 18.92 7 650 40.16Endopeptidase O OS =- Streptococcus Intracellular pneumoniae (strainCGSP14) GN = pepO PE = 4 SV = 1 - [B2IRQ3_STRPS] B2IR80 26.03 6 29239.76 Uncharacterized protein OS = Multi- Streptococcus pneumoniae(strain transmembrane CGSP14) GN = SPCG_1547 PE = 4 SV = 1 -[B2IR80_STRPS] B2INZ2 20.82 6 413 39.4 D-alanyl-D-alanine N-carboxypeptidase OS =- Streptococcus terminally pneumoniae (strainCGSP14) anchored GN = dacA PE = 3 SV = 1 - (with CS) [B2INZ2_STRPS]B2IR93 45.95 6 259 39.26 Triosephosphate isomerase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = tpiA PE = 3 SV = 1 -[B2IR93_STRPS] B2IP56 34.48 8 290 39.23 Uncharacterized protein OS =Lipid Streptococcus pneumoniae (strain anchored CGSP14) GN = SPCG_0875PE = 4 SV = 1 - [B2IP56_STRPS] B2IPZ7 46.59 6 279 39.17 DegV familyprotein OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1169 PE = 4 SV = 1 - [B2IPZ7_STRPS] B2INK5 19.52 8 502 39.03Glycerol kinase OS =- Streptococcus Intracellular pneumoniae (strainCGSP14) GN = glpK PE = 3 SV = 1 - [GLPK_STRPS] B2INM2 32.67 7 150 38.8450S ribosomal protein L9 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = rplI PE = 3 SV = 1 - [RL9_STRPS] B2IPH6 21.17 4 35938.78 Peptide chain release factor 1 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = prfA PE = 3 SV = 1 - [RF1_STRPS] B2ING044.21 3 95 38.52 30S ribosomal protein S16 OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = rpsP PE = 3 SV = 1 -[RS16_STRPS] B2IQZ0 33.07 7 378 38.34 DNA polymerase III subunit betaIntracellular OS =- Streptococcus pneumoniae (strain CGSP14) GN = dnaNPE = 4 SV = 1 - [B2IQZO_STRPS] B2IR11 29.29 8 420 37.79 DNA repairprotein radA OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = radA PE = 3 SV = 1 - [B2IR11_STRPS] B2ISK0 7.86 8 1463 37.3 DNApolymerase III PolC-type OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = polC PE = 3 SV = 1 - [B2ISKO_STRPS] B2IMD8 40.15 8264 37.04 Uncharacterized protein OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0513 PE = 4 SV = 1 - [B2IMD8_STRPS]B2IP06 33.66 7 303 36.91 Tagatose-6-phosphate kinase OS IntracellularStreptococcus pneumoniae (strain CGSP14) GN = fruB PE = 3 SV = 1 -[B2IP06_STRPS] B2IPT6 46.2 5 171 36.86 Galactose-6-phosphate isomeraseIntracellular subunit LacB OS =- Streptococcus pneumoniae (strainCGSP14) GN = lacB PE = 3 SV = 1 - [LACB_STRPS] B2IMT1 28.33 7 406 36.74Beta-lactam resistance factor OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = murM PE = 4 SV = 1 - [B2IMT1_STRPS]B2IN52 24.87 8 567 36.72 Dihydroxy-acid dehydratase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = ilvD PE = 3 SV = 1 -[ILVD_STRPS] B2IRH4 23.13 5 320 36.59 Uncharacterized protein OS = N-Streptococcus pneumoniae (strain terminally CGSP14) GN = SPCG_0093 PE =4 SV = 1 - anchored [B2IRH4_STRPS] (No CS) B2IME1 27.51 7 378 36.3Transcription Intracellular termination/antitermination protein NusA OS=- Streptococcus pneumoniae (strain CGSP14) GN = nusA PE = 3 SV = 1 -[B2IME1 STRPS] B2ISX2 51.67 9 240 36.23 Pseudouridine synthase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rluB PE = 3SV = 1 - [B2ISX2_STRPS] B2IN61 77.55 6 49 35.89 50S ribosomal proteinL33 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =rpmG PE = 3 SV = 1 - [RL33_STRPS] B2IQ98 60.64 4 94 35.66 50S ribosomalprotein L31 type B Intracellular OS =- Streptococcus pneumoniae (strainCGSP14) GN = rpmE PE = 3 SV = 1 - [B2IQ98_STRPS] B2IPY5 32.19 7 37935.53 Glycogen biosynthesis protein GlgD Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = glgD PE = 4 SV = 1 -[B2IPY5_STRPS] B2IRA3 21.23 4 438 35.35 Pyridine nucleotide-disulfideIntracellular oxidoreductase OS =- Streptococcus pneumoniae (strainCGSP14) GN = merA PE = 3 SV = 1 - [B2IRA3_STRPS] B2IR65 48.8 6 209 35.25Uncharacterized protein OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1532 PE = 4 SV = 1 - [B2IR65_STRPS] B2INN136.99 7 346 34.94 Elongation factor Ts OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = tsf PE = 3 SV = 1 - [EFTS_STRPS] B2IRV519.83 3 343 34.85 Phospho-2-dehydro-3- Intracellular deoxyheptonatealdolase OS = Streptococcus pneumoniae (strain CGSP14) GN = aroF PE = 3SV = 1 - [B2IRV5_STRPS] B2IN60 53.33 4 60 34.75 50S ribosomal proteinL32 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =rpmF PE = 3 SV = 1 - [RL32_STRPS] B2IQ13 32.08 6 371 34.73Aminotransferase, class-V OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1185 PE = 3 SV = 1 - [B2IQ13_STRPS] B2INQ136.11 5 180 34.11 Transcriptional regulator, TetR Intracellular familyOS =- Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2200 PE = 4 SV= 1 - [B2INQ1_STRPS] B2IP08 15.91 7 767 33.57 SpoE family protein OS =Multi- Streptococcus pneumoniae (strain transmembrane CGSP14) GN = ftsKPE = 3 SV = 1 - [B2IP08_STRPS] B2IRW8 41.24 6 177 33.56 Transcriptionalrepressor NrdR OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = nrdR PE = 3 SV = 1 - [B2IRW8_STRPS] B2ISL9 57.61 6 18433.49 GTP cyclohydrolase 1 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = foIE PE = 3 SV = 1 - [GCH1_STRPS] B2IQL2 25.35 4217 32.6 Phosphate-specific transport Intracellular system accessoryprotein PhoU OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1384 PE = 3 SV = 1 - [B2IQL2_STRPS] B2ILW1 39.58 4 144 32.47Transcriptional regulator, MarR Intracellular family OS =- Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0414 PE = 4 SV = 1 - [B2ILW1_STRPS]B2IS33 15.25 5 518 32.44 Competence-induced protein Ccs4 Multi- OS =-Streptococcus pneumoniae (strain transmembrane CGSP14) GN = SPCG_0211 PE= 4 SV = 1 - [B2IS33_STRPS] B2INX7 16.53 4 236 32.43 Uncharacterizedprotein OS = Multi- Streptococcus pneumoniae (strain transmembraneCGSP14) GN = SPCG_0802 PE = 4 SV = 1 - [B2INX7_STRPS] B2INM3 14.16 7 65732.29 DHH subfamily 1 protein OS = Multi- Streptococcus pneumoniae(strain transmembrane CGSP14) GN = SPCG_2172 PE = 4 SV = 1 -[B2INM3_STRPS] B2INY8 22.62 6 420 32.18 Uncharacterized protein OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0813 PE= 4 SV = 1 - [B2INY8_STRPS] B2IN02 20.77 5 443 32.03 Sensor histidinekinase PnpS OS = Multi- Streptococcus pneumoniae (strain transmembraneCGSP14) GN = SPCG_2049 PE = 4 SV = 1 - [B2IN02_STRPS] B2IMR8 13.73 5 45931.87 Transmembrane protein Vexp3 OS = Multi- Streptococcus pneumoniae(strain transmembrane CGSP14) GN = SPCG_0562 PE = 3 SV = 1 -[B2IMR8_STRPS] B2IP11 34.65 7 404 31.83 Probable tRNA sulfurtransferaseIntracellular OS =- Streptococcus pneumoniae (strain CGSP14) GN = thiIPE = 3 SV = 1 - [THII_STRPS] B2INZ1 26.38 8 470 31.82 FeS assemblyprotein SufB OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = SPCG_0816 PE = 4 SV = 1 - [B2INZ1_STRPS] B2IQ37 41.27 5 252 31.78ABC transporter, ATP-binding Intracellular protein OS =- Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1209 PE = 4 SV = 1 - [B2IQ37_STRPS]B2IN78 31.83 6 399 31.49 Cell division protein DivIB OS = Intracellular/Streptococcus pneumoniae (strain TMH start CGSP14) GN = divIB PE = 3 SV= 1 - AFTER 60 [B2IN78_STRPS] B2IS71 21.69 5 249 31.41 Phosphoglyceratemutase family Intracellular protein OS =- Streptococcus pneumoniae(strain CGSP14) GN = gpmB PE = 4 SV = 1 - [B2IS71_STRPS] B2IQJ2 23.71 5388 31.39 Chorismate synthase OS = = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = aroC PE = 3 SV = 1 - [AROC_STRPS] B2IST913.89 7 583 31.29 ABC transporter, ATP- Multi- binding/permease proteinOS = transmembrane Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1814 PE = 4 SV = 1 - [B2IST9_STRPS] B2IPQ2 32.05 6 234 30.92Uncharacterized protein OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1072 PE = 4 SV = 1 - [B2IPQ2_STRPS] B2ISM018.89 3 270 30.69 7,8-dihydroneopterin aldolase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = suID PE = 3 SV = 1 -[B2ISMO_STRPS] B2IMH1 16.74 6 472 30.67 Cof family protein OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = SPCG_1962 PE= 4 SV = 1 - [B2IMH1_STRPS] B2ISV2 17.09 5 392 30.34 Galactokinase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = gaIK PE = 3SV = 1 - [B2ISV2_STRPS] B2IR20 12.6 8 889 30.19 DNA polymerase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = poIA PE = 3SV = 1 - [B2IR20_STRPS] B2IR50 26.4 6 481 30UDP-N-acetylmuramoyl-L-alanyl-D- Intracellular glutamate--L-lysineligase OS = Streptococcus pneumoniae (strain CGSP14) GN = murE PE = 3 SV= 1 - [B2IR50_STRPS] B2ISI6 20.93 6 258 29.94 Isoprenyl transferase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = uppS PE = 3SV = 1 - [B2IS16_STRPS] B2IMQ1 31.47 7 375 29.85 Phenylalanine--tRNAligase alpha Intracellular subunit OS =- Streptococcus pneumoniae(strain CGSP14) GN = pheS PE = 3 SV = 1 - [B2IMQ1_STRPS] B2ISS1 20.39 6407 29.77 Tryptophan synthase beta chain Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = trpB PE = 3 SV = 1 -[TRPB_STRPS] B2IMZ0 22.05 5 449 29.65 Glucose-6-phosphate isomeraseIntracellular OS =- Streptococcus pneumoniae (strain CGSP14) GN = pgi PE= 3 SV = 1 - [G6PI_STRPS] B2IRD1 13.75 4 778 29.59 Cation-transportingATPase, E1-E2 Multi- family OS =- Streptococcus pneumoniae transmembrane(strain CGSP14) GN = ctpE PE = 3 SV = 1 - [B2IRD1_STRPS] B2IQ17 51.09 5229 29.49 Glutamine amidotransferase, class I Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1189 PE = 4 SV = 1 -[B2IQ17_STRPS] B2IPV5 30.5 5 200 29.44 Uncharacterized protein OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1127 PE= 4 SV = 1 - [B2IPV5_STRPS] B2ILW5 27.78 6 306 29.34 Malonyl CoA-acylcarrier protein Intracellular transacylase OS =- Streptococcuspneumoniae (strain CGSP14) GN = fabD PE = 3 SV = 1 - [B2ILW5_STRPS]B2IQ16 29.48 6 424 29.26 Psr protein OS =- Streptococcus Intracellular/pneumoniae (strain CGSP14) TMH start GN = SPCG_1358 PE = 4 SV = 1 -AFTER 60 [B2IQ16_STRPS] B2IPB3 40.51 5 195 28.99 Translation initiationfactor IF-3 OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = infC PE = 3 SV = 1 - [B2IPB3_STRPS] B2IM63 18.52 7 432 28.92Competence-induced protein Ccs50 N- OS =- Streptococcus pneumoniae(strain terminally CGSP14) GN = SPCG_1944 PE = 4 SV = 1 - anchored[B2IM63_STRPS] (No CS) B2IP99 44.53 6 247 28.66 Uridylate kinase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = pyrH PE = 3SV = 1 - [B2IP99_STRPS] B2IPF5 10.79 5 834 28.65 Uncharacterized proteinOS = N- Streptococcus pneumoniae (strain terminally CGSP14) GN =SPCG_0977 PE = 4 SV = 1 - anchored [B2IPF5_STRPS] (No CS) B2IM26 26.1 6318 28.61 Autolysin OS =- Streptococcus Intracellular pneumoniae (strainCGSP14) GN = lytA PE = 4 SV = 1 - [B2IM26_STRPS] B2INF6 43.1 5 239 28.52tRNA (guanine-N(1)-)- Intracellular methyltransferase OS =-Streptococcus pneumoniae (strain CGSP14) GN = trmD PE = 3 SV = 1 -[TRMD_STRPS] B2ISI0 15.25 6 833 28.41 Leucine--tRNA ligase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = leuS PE = 3SV = 1 - [SYL_STRPS] B2IQH8 20.5 6 439 28.08 Homoserine dehydrogenase OS= Intracellular Streptococcus pneumoniae (strain CGSP14) GN = hom PE = 3SV = 1 - [B2IQH8_STRPS] B2INT8 57.89 3 76 27.91 Uncharacterized proteinOS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0763 PE = 4 SV = 1 - [B2INT8_STRPS] B2IMD9 15.64 2 211 27.64 tRNA(guanine-N(7)-)- Intracellular methyltransferase OS =- Streptococcuspneumoniae (strain CGSP14) GN = trmB PE = 3 SV = 1 - [TRMB_STRPS] B2IQS226.88 4 253 27.63 SpoU rRNA methylase family Intracellular protein OS =-Streptococcus pneumoniae (strain CGSP14) GN = trmH PE = 3 SV = 1 -[B2IQS2_STRPS] B2IR73 16.69 5 623 27.4 ABC transporter, ATP-bindingIntracellular protein OS =- Streptococcus pneumoniae (strain CGSP14) GN= SPCG_1540 PE = 4 SV = 1 - [B2IR73_STRPS] B2IPP4 10.12 6 662 27.29UvrABC system protein B OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = uvrB PE = 3 SV = 1 - [UVRB_STRPS] B2IQK6 26.91 4301 27.27 UDP-N- Intracellular acetylenolpyruvoylglucosamine reductaseOS =- Streptococcus pneumoniae (strain CGSP14) GN = murB PE = 3 SV = 1 -[MURB_STRPS] B2IP98 25.45 6 444 27.26 Methylenetetrahydrofolate--tRNA-Intracellular (uracil-5-)-methyltransferase TrmFO OS =- Streptococcuspneumoniae (strain CGSP14) GN = trmFO PE = 3 SV = 2 - [TRMFO _STRPS]B2IQK5 19.48 5 385 27.06 Spermidine/putrescine import ATP- Intracellularbinding protein PotA OS = Streptococcus pneumoniae (strain CGSP14) GN =potA PE = 3 SV = 1 - [B2IQK5_STRPS] B2INN3 30.61 7 392 26.86 Secreted 45kd protein OS = Secretory Streptococcus pneumoniae (strain (released)CGSP14) GN = SPCG_2182 PE = 4 SV = 1 - (with CS) [B2INN3_STRPS] B2IN2040.78 4 103 26.71 Uncharacterized protein OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = SPCG_2067 PE = 4 SV = 1 -[B2IN20_STRPS] B2IPN6 8.14 6 1179 26.7 Chromosome partition protein SmcIntracellular OS =- Streptococcus pneumoniae (strain CGSP14) GN = smc PE= 3 SV = 1 - [B2IPN6_STRPS] B2IRW9 52.07 5 121 26.67 Transcriptionalregulator, GntR Intracellular family OS =- Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1686 PE = 4 SV = 1 - [B2IRW9_STRPS] B2IS8413.47 2 438 26.55 Ribosomal RNA small subunit Intracellularmethyltransferase B OS = Streptococcus pneumoniae (strain CGSP14) GN =sunL PE = 3 SV = 1 - [B2IS84_STRPS] B2IMX9 19.88 7 332 26.49Pneumococcal surface protein, Lipid putative OS =- Streptococcusanchored pneumoniae (strain CGSP14) GN = SPCG_0623 PE = 4 SV = 1 -[B2IMX9_STRPS] B2IP15 8.69 4 541 26.36 RNA methyltransferase, TrmAIntracellular family OS =- Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1007 PE = 3 SV = 1 - [B2IPI5_STRPS] B2IPS8 15.14 7 555 25.77 DNArepair protein RecN OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = recN PE = 3 SV = 1 - [B2IPS8_STRPS] B2INY5 17.06 6 55125.65 DNA polymerase III subunits Intracellular gamma and tau OS =-Streptococcus pneumoniae (strain CGSP14) GN = dnaX PE = 4 SV = 1 -[B2INY5_STRPS] B2ISU0 10.1 4 594 25.61 ABC transporter, ATP- Multi-binding/permease protein OS = transmembrane Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1815 PE = 4 SV = 1 - [B2ISUO_STRPS] B2INU624.53 6 424 25.32 Phosphopentomutase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = deoB PE = 3 SV = 1 - [B2INU6_STRPS]B2IM77 17.44 2 281 25.26 Undecaprenyl-diphosphatase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = uppP PE = 3 SV = 1 -[UPPP_STRPS] B2IS76 21.11 5 398 25.26 Hydroxymethylglutaryl-CoAIntracellular synthase OS =- Streptococcus pneumoniae (strain CGSP14) GN= mvaS PE = 4 SV = 1 - [B2IS76_STRPS] B2IPD9 20.3 4 266 24.91Uncharacterized protein OS = N- Streptococcus pneumoniae (strainterminally CGSP14) GN = SPCG_0961 PE = 4 SV = 1 - anchored[B2IPD9_STRPS] (No CS) B2IP88 14.52 6 420 24.84 Gamma-glutamyl phosphateIntracellular reductase OS =- Streptococcus pneumoniae (strain CGSP14)GN = proA PE = 3 SV = 1 - [PROA_STRPS] B2IQU2 18.36 4 305 24.77Glycine--tRNA ligase alpha subunit Intracellular OS =- Streptococcuspneumoniae (strain CGSP14) GN = glyQ PE = 3 SV = 1 - [SYGA_STRPS] B2INW025 5 220 24.67 Deoxyribose-phosphate aldolase Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = deoC PE = 3 SV = 1 -[DEOC_STRPS] B2IRB6 28.25 6 223 24.63 Cytidylate kinase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = cmk PE = 3SV = 1 - [KCY_STRPS] B2IP93 24.91 5 289 24.4 Ribosomal RNA small subunitIntracellular methyltransferase I OS = Streptococcus pneumoniae (strainCGSP14) GN = rsml PE = 3 SV = 1 - [B2IP93_STRPS] B2IPN4 29.88 5 32824.28 GMP reductase OS =- Streptococcus Intracellular pneumoniae (strainCGSP14) GN = guaC PE = 3 SV = 1 - [GUAC_STRPS] B2IN17 24.9 5 257 24.1Fucose operon repressor, putative Intracellular OS =- Streptococcuspneumoniae (strain CGSP14) GN = fcsR PE = 4 SV = 1 - [B2IN17_STRPS]B21MP1 31.07 5 206 24.09 Hydrolase, haloacid dehalogenase- Intracellularlike family OS =- Streptococcus pneumoniae (strain CGSP14) GN =SPCG_2031 PE = 4 SV = 1 - [B2IMP1_STRPS] B2IRV6 17.78 3 343 23.78Phospho-2-dehydro-3- Intracellular deoxyheptonate aldolase OS =Streptococcus pneumoniae (strain CGSP14) GN = aroG PE = 3 SV = 1 -[B2IRV6_STRPS] B2IQ68 27.02 5 359 23.7 Carbamoyl-phosphate synthaseIntracellular small chain OS =- Streptococcus pneumoniae (strain CGSP14)GN = carA PE = 3 SV = 1 - [B2IQ68_STRPS] B2IMY1 17.2 3 279 23.45Thymidylate synthase OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = thyA PE = 3 SV = 1 - [B2IMY1_STRPS] B2IM44 17.88 3 27423.39 DNA-entry nuclease OS = N- Streptococcus pneumoniae (strainterminally CGSP14) GN = endA PE = 4 SV = 1 - anchored [B2IM44_STRPS] (NoCS) B2IR18 22.54 3 213 23.34 Amino acid ABC transporter, ATP-Intracellular binding protein, putative OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_0107 PE = 4 SV = 1 - [B2IRI8_STRPS] B2IRQ15.51 2 653 23.17 PTS system IIABC components OS = Multi- Streptococcuspneumoniae (strain transmembrane CGSP14) GN = scrA PE = 4 SV = 1 -[B2IRQ1_STRPS] B2IQR5 19.92 4 246 23.16 CppA protein OS =- StreptococcusIntracellular pneumoniae (strain CGSP14) GN = cppA PE = 4 SV = 1 -[B2IQR5_STRPS] B2ILR4 22.22 2 117 23.07 Cell cycle protein GpsB OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = gpsB PE = 3SV = 1 - [B2ILR4_STRPS] B2IQN2 20.25 4 316 22.86 HPrkinase/phosphorylase OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = hprK PE = 3 SV = 1 - [B2IQN2_STRPS] B2IMA5 25 6 448 22.78Glutamine synthetase, type I OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = glnA PE = 3 SV = 1 - [B2IMA5_STRPS] B2IQ75 33.87 4186 22.38 LemA protein OS =- Streptococcus N- pneumoniae (strain CGSP14)terminally GN = lemA PE = 4 SV = 1 - anchored [B2IQ75_STRPS] (No CS)B2INX5 19.41 5 340 21.91 Branched-chain-amino-acid Intracellularaminotransferase OS =- Streptococcus pneumoniae (strain CGSP14) GN =ilvE PE = 3 SV = 1 - [B2INX5_STRPS] B2IMY7 43.48 4 207 21.8Uncharacterized protein OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = SPCG_0630 PE = 4 SV = 1 - [B2IMY7_STRPS] B2IPY614.21 4 380 21.63 Glucose-1-phosphate Intracellular adenylyltransferaseOS = Streptococcus pneumoniae (strain CGSP14) GN = glgC PE = 3 SV = 1 -[GLGC_STRPS] B2IM69 20.91 4 416 21.42 L-threonine dehydratase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = ilVA PE = 3SV = 1 - [B2IM69_STRPS] B2INA8 35.36 5 263 21.29 Phosphomethylpyrimidinekinase Intracellular OS =- Streptococcus pneumoniae (strain CGSP14) GN =thiD PE = 4 SV = 1 - [B2INA8_STRPS] B2ISK9 13.48 3 267 21.26 PTS system,mannose-specific IIC Multi- component OS =- Streptococcustransmembrane(Lipid pneumoniae (strain CGSP14) modified GN = manM PE = 4SV = 1 - N-termini) [B2ISK9_STRPS] B2IQ32 12.93 3 441 21.17 Glycosyltransferase, group 1 OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1204 PE = 4 SV = 1 - [B2IQ32_STRPS] B21M58 8.12 3 30820.91 Membrane protein insertase YidC Multi- OS =- Streptococcuspneumoniae (strain transmembrane CGSP14) GN = yidC PE = 3 SV = 1 -[B2IM58_STRPS] B2INR5 26.54 4 309 20.79 Oxidoreductase, aldo/ketoIntracellular reductase family OS =- Streptococcus pneumoniae (strainCGSP14) GN = SPCG_0740 PE = 4 SV = 1 - [B2INR5_STRPS] B2INC9 36.22 3 19620.35 ATP-dependent Clp protease Intracellular proteolytic subunit OS =CGSP14) GN = clpP PE = 3 SV = 1 - [CLPP_STRPS] B2IQ35 25.75 4 369 20.25RNA polymerase sigma factor SigA Intracellular OS =- Streptococcuspneumoniae (strain CGSP14) GN = rpoD PE = 3 SV = 1 - [B2IQ35_STRPS]B21MR7 20 3 215 20.15 ABC transporter, ATP-binding Intracellular proteinVexp2 OS =- Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0561 PE =4 SV = 1 - [B2IMR7_STRPS]

TABLE 6 Proteins identified in MP from serotype 3. The accession numbersrefer to SwissProt on the date of filing. Accession # # CoveragePeptides #AAs Score Description Localization B2IQV2 90.84 45 404 2064.28Elongation factor Tu OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = tuf PE = 3 SV = 1 - [B2IQV2_STRPS] B2IRT9 69.68 49 4421423.4 Sugar ABC transporter, sugar- Lipid binding protein OS =-Streptococcus anchored pneumoniae (strain CGSP14) GN = SPCG_1656 PE = 4SV = 1 - [B2IRT9_STRPS] B2INY1 82.25 60 400 1396.37 30S ribosomalprotein S1 OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = rpsA PE = 4 SV = 1 - [B2INY1_STRPS] B2ISJ9 86.15 57 693 1352.29Elongation factor G OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = fusA PE = 3 SV = 1 - [EFG_STRPS] B2IPX8 86.64 41 4341214.93 Enolase OS = -Streptococcus Intracellular pneumoniae (strainCGSP14) GN = eno PE = 3 SV = 1 - [ENO_STRPS] B2INB2 75.63 49 591 1080.64Pyruvate oxidase OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = spxB PE = 3 SV = 1 - [B2INB2_STRPS] B2IN25 76.44 47 4501068.79 Maltose/maltodextrin ABC Lipid transporter,maltose/maltodextrin- anchored binding protein OS = -Streptococcuspneumoniae (strain CGSP14) GN = SPCG_2072 PE = 4 SV = 1 - [B2IN25_STRPS]B2INK4 76.15 51 608 829.85 Uncharacterized protein OS = N- Streptococcuspneumoniae (strain terminally CGSP14) GN = SPCG_2153 PE = 4 anchored SV= 1 - [B2INK4_STRPS] (No CS) B2ISK8 52.48 17 303 715.49 PTS system,mannose-specific IID Multi- component OS = -Streptococcus transmembranepneumoniae (strain CGSP14) GN = manN PE = 4 SV = 1 - [B2ISK8_STRPS]B2IR98 86.7 33 376 633.37 Sugar ABC transporter, ATP-bindingIntracellular protein OS = -Streptococcus pneumoniae (strain CGSP14) GN= SPCG_1565 PE = 3 SV = 1 - [B2IR98_STRPS] B2IMI7 75.21 29 359 616.73Glyceraldehyde-3-phosphate Intracellular dehydrogenase OS =-Streptococcus pneumoniae (strain CGSP14) GN = gapA PE = 3 SV = 1 -[B2IMI7_STRPS] B2ISL0 55.12 20 332 608.4 PTS system, mannose-specificIIAB Intracellular components OS = -Streptococcus pneumoniae (strainCGSP14) GN = manL PE = 4 SV = 1 - [B2ISL0_STRPS] B2IRH0 78.91 39 512573.11 ABC transporter, substrate-binding Lipid protein OS =-Streptococcus anchored pneumoniae (strain CGSP14) GN = SPCG_0089 PE = 4SV = 1 - [B2IRH0_STRPS] B2IS43 74.01 23 277 570.24 50S ribosomal proteinL2 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpIBPE = 3 SV = 1 - [RL2_STRPS] B2INK5 70.12 34 502 534.8 Glycerol kinase OS= -Streptococcus Intracellular pneumoniae (strain CGSP14) GN = glpK PE =3 SV = 1 - [GLPK_STRPS] B2IRV7 73.6 55 837 530.96 Protein translocasesubunit SecA Intracellular OS = -Streptococcus pneumoniae (strainCGSP14) GN = secA PE = 3 SV = 1 - [B2IRV7_STRPS] B2INW2 65.51 28 374519.89 Lipoprotein OS = -Streptococcus Lipid pneumoniae (strain CGSP14)anchored GN = SPCG_0787 PE = 4 SV = 1 - [B2INW2_STRPS] B2IPD4 58.15 28313 480.11 Foldase protein PrsA OS = Lipid Streptococcus pneumoniae(strain anchored CGSP14) GN = prsA PE = 3 SV = 1 - [PRSA_STRPS] B2IR0059.97 38 652 462.58 ATP-dependent zinc Multi- metalloprotease FtsH OS =transmembrane Streptococcus pneumoniae (strain CGSP14) GN = ftsH PE = 3SV = 1 - [B2IR00_STRPS] B2IM39 54.12 57 1225 460.95 DNA-directed RNApolymerase Intracellular subunit beta' OS = -Streptococcus pneumoniae(strain CGSP14) GN = rpoC PE = 3 SV = 1 - [RPOC_STRPS] B2IM81 63.18 37774 432.81 Formate acetyltransferase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = pfl PE = 4 SV = 1 - [B2IM81_STRPS]B2IM49 72.73 25 330 394.5 Aspartate--ammonia ligase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = asnA PE = 3 SV = 1 -[ASNA_STRPS] B2IN96 69.05 25 378 394.18 Lactate oxidase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = IctO PE = 4SV = 1 - [B2IN96_STRPS] B2IPT8 93.56 27 326 382.46 Tagatose1,6-diphosphate aldolase Intracellular OS = -Streptococcus pneumoniae(strain CGSP14) GN = lacD PE = 3 SV = 1 - [B2IPT8_STRPS] B2INN2 73.36 26259 380.63 30S ribosomal protein S2 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = rpsB PE = 3 SV = 2 - [RS2_STRPS] B2IQX086.32 28 468 375.02 ATP synthase subunit beta OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = atpD PE = 3 SV = 1 -[ATPB_STRPS] B2IMA2 85.18 25 398 374.17 Phosphoglycerate kinase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = pgk PE = 3SV = 1 - [PGK_STRPS] B2IS46 81.11 26 217 368.3 30S ribosomal protein S3OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpsC PE= 3 SV = 1 - [RS3_STRPS] B2ISZ0 64.49 31 659 336.33 Oligopeptide ABCtransporter, Lipid oligopeptide-binding protein AmiA anchored OS =-Streptococcus pneumoniae (strain CGSP14) GN = amiA PE = 4 SV = 1 -[B2ISZ0_STRPS] B2ISG3 77.23 21 202 332.38 General stress protein 24,putative Intracellular OS = -Streptococcus pneumoniae (strain CGSP14) GN= SPCG_1781 PE = 4 SV = 1 - [B2ISG3_STRPS] B21ML8 69.46 25 406 332.33Acetate kinase OS = -Streptococcus Intracellular pneumoniae (strainCGSP14) GN = ackA PE = 3 SV = 1 - [B2IML8_STRPS] B2IP54 71.66 27 501328.6 Pyruvate kinase OS = -Streptococcus Intracellular pneumoniae(strain CGSP14) GN = SPCG_0873 PE = 3 SV = 1 - [B2IP54_STRPS] B2IN1057.69 12 338 324.09 Glycerol-3-phosphate Intracellular dehydrogenase[NAD(P)+] OS = Streptococcus pneumoniae (strain CGSP14) GN = gpsA PE = 3SV = 1 - [GPDA_STRPS] B2IRR1 66.09 12 230 318.072,3-bisphosphoglycerate- Intracellular dependent phosphoglycerate mutaseOS = -Streptococcus pneumoniae (strain CGSP14) GN = gpmA PE = 3 SV = 1 -[GPMA_STRPS] B2IRI6 36.84 5 95 310.42 Bacteriocin, putative OS = N-terminally Streptococcus pneumoniae (strain anchored CGSP14) GN =SPCG_0105 PE = 4 (with CS) SV = 1 - [B2IRI6_STRPS] B2IQY1 57.35 26 551302.6 Uncharacterized protein OS = Intracellular/ Streptococcuspneumoniae (strain TMH CGSP14) GN = SPCG_1503 PE = 4 SV = 1 - start[B2IQY1_STRPS] AFTER 60 B2ISJ7 67.88 14 137 301.47 30S ribosomal proteinS12 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =rpsL PE = 3 SV = 1 - [RS12_STRPS] B2IQX2 38.92 21 501 285.3 ATP synthasesubunit alpha OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = atpA PE = 3 SV = 1 - [ATPA_STRPS] B2IP53 50.15 16 335269.91 ATP-dependent 6- Intracellular phosphofructokinase OS =Streptococcus pneumoniae (strain CGSP14) GN = pfkA PE = 3 SV = 1 -[PFKA_STRPS] B2IR47 51.69 27 652 263.77 Oligopeptide ABC transporter,Lipid oligopeptide-binding protein AliB anchored OS = -Streptococcuspneumoniae (strain CGSP14) GN = aliB PE = 4 SV = 1 - [B2IR47_STRPS]B2ISZ6 50.36 17 419 260.6 Sugar ABC transporter, sugar- Lipid bindingprotein OS = -Streptococcus anchored pneumoniae (strain CGSP14) GN =SPCG_1871 PE = 4 SV = 1 - [B2ISZ6_STRPS] B2IMB5 52.72 24 607 254.75Chaperone protein Dnak OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = dnaK PE = 3 SV = 1 - [DNAK_STRPS] B2INT1 42.12 26584 253.89 Septation ring formation regulator N- EzrA OS =-Streptococcus pneumoniae terminally (strain CGSP14) GN = ezrA PE = 3 SV= 1 - anchored [B2INT1_STRPS] (No CS) B2IM12 66.03 26 471 237.78Pneumolysin OS = -Streptococcus Intracellular pneumoniae (strain CGSP14)GN = ply PE = 4 SV = 1 - [B2IM12_STRPS] B2IS47 55.47 11 137 228.66 50Sribosomal protein L16 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = rpIP PE = 3 SV = 1 - [RL16_STRPS] B2IRM9 68.48 18276 218.21 ABC transporter, substrate-binding Lipid protein OS =-Streptococcus anchored pneumoniae (strain CGSP14) GN = SPCG_0152 PE = 4SV = 1 - [B2IRM9_STRPS] B2IRG4 64.53 23 203 217.88 30S ribosomal proteinS4 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpsDPE = 3 SV = 1 - [RS4_STRPS] B2IS57 86.59 19 164 213.33 30S ribosomalprotein S5 OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = rpsE PE = 3 SV = 1 - [B2IS57_STRPS] B2IM40 44.9 29.1 4 824 208.41DNA-directed RNA polymerase Intracellular subunit beta OS =-Streptococcus pneumoniae (strain CGSP14) GN = rpoB PE = 3 SV = 1 -[B2IM40_STRPS] B2ISM3 87.16 20 148 207.33 50S ribosomal protein L13 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpIM PE = 3SV = 1 - [B2ISM3_STRPS] B2IQ88 76.52 13 115 207.04 50S ribosomal proteinL19 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =rpIS PE = 3 SV = 1 - [RL19_STRPS] B2ISD1 80.77 12 104 201.22 ThioredoxinOS = -Streptococcus Intracellular pneumoniae (strain CGSP14) GN = trxAPE = 3 SV = 1 - [B2ISD1_STRPS] B2IS40 62.98 15 208 197.25 50S ribosomalprotein L3 OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = rpIC PE = 3 SV = 1 - [RL3_STRPS] B2IMJ9 41.57 28 890 195.17Aldehyde-alcohol dehydrogenase Intracellular OS = -Streptococcuspneumoniae (strain CGSP14) GN = adhE PE = 3 SV = 1 - [B2IMJ9_STRPS]B2IPW4 45.86 24 567 191.95 Dihydrolipoyl dehydrogenase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = acoL PE = 4SV = 1 - [B2IPW4_STRPS] B2IP07 28.31 14 650 190.79 PTS system, fructosespecific IIABC Multi- components OS = -Streptococcus transmembranepneumoniae (strain CGSP14) GN = fruA PE = 4 SV = 1 - [B2IP07_STRPS]B2IS55 84.27 13 178 180.96 50S ribosomal protein L6 OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = rpIF PE = 3 SV = 1 -[RL6_STRPS] B2IME4 33.44 20 930 180.14 Translation initiation factorIF-2 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =infB PE = 3 SV = 1 - [IF2_STRPS] B2ISW0 53.72 12 242 178.66 Cholinetransporter OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = SPCG_1835 PE = 4 SV = 1 - [B2ISW0_STRPS] B2IS66 43.31 8 127 176.9830S ribosomal protein S11 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = rpsK PE = 3 SV = 1 - [RS11_STRPS] B2IPR1 43.98 13332 173.61 L-lactate dehydrogenase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = Idh PE = 3 SV = 1 - [B2IPR1_STRPS]B2IRS1 44.15 15 419 169.63 Cell division protein FtsZ OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = ftsZ PE = 3 SV = 1 -[B2IRS1_STRPS] B2IPP2 36.89 22 721 166.14 Amino acid ABC transporter,amino Multi- acid-binding protein/permease transmembrane protein OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1062 PE = 3 SV = 1 -[B2IPP2_STRPS] B2ILZ5 43.89 21 540 165.77 60 kDa chaperonin OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = groL PE = 3SV = 1 - [CH60_STRPS] B2IS41 65.22 16 207 162.59 50S ribosomal proteinL4 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpIDPE = 3 SV = 1 - [RL4_STRPS] B2ILW6 75.31 15 243 162.563-ketoacyl-(Acyl-carrier-protein) Intracellular reductase OS =-Streptococcus pneumoniae (strain CGSP14) GN = fabG PE = 4 SV = 1 -[B2ILW6_STRPS] B2ISY9 34.54 14 498 161.94 Oligopeptide ABC transporter,Multi- permease protein Amic OS = transmembrane Streptococcus pneumoniae(strain CGSP14) GN = amiC PE = 3 SV = 1 - [B2ISY9_STRPS] B2ISY6 58.31 18331 161.56 Oligopeptide ABC transporter, ATP- Intracellular bindingprotein AmiF OS = Streptococcus pneumoniae (strain CGSP14) GN = amiF PE= 3 SV = 1 - [B2ISY6_STRPS] B2IS52 77.22 13 180 159.07 50S ribosomalprotein L5 OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = rplE PE = 3 SV = 1 - [RL5_STRPS] B2ILW7 50.24 10 414 153.033-oxoacyl-[acyl-carrier-protein] Intracellular synthase 2 OS =-Streptococcus pneumoniae (strain CGSP14) GN = fabF PE = 3 SV = 1 -[B2ILW7_STRPS] B2ISY7 46.76 16 355 151.21 Oligopeptide ABC transporter,ATP- Intracellular binding protein AmiE OS = Streptococcus pneumoniae(strain CGSP14) GN = amiE PE = 3 SV = 1 - [B2ISY7_STRPS] B2ILU5 34.89 13427 150.71 Trigger factor OS = -Streptococcus Intracellular pneumoniae(strain CGSP14) GN = tig PE = 3 SV = 1 - [TIG_STRPS] B2IMU8 55.46 15 229148.33 50S ribosomal protein L1 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = rpIA PE = 3 SV = 1 - [RL1_STRPS] B2IS4456.99 10 93 148.01 30S ribosomal protein S19 OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = rpsS PE = 3 SV = 1 -[RS19_STRPS] B2IQT6 47.49 18 459 146.83 NADH oxidase OS = -StreptococcusIntracellular pneumoniae (strain CGSP14) GN = nox PE = 4 SV = 1 -[B2IQT6_STRPS] B2INP5 40.65 15 492 145.57 Inosine-5′-monophosphateIntracellular dehydrogenase OS = -Streptococcus pneumoniae (strainCGSP14) GN = imdH PE = 3 SV = 1 - [B2INP5_STRPS] B2INF2 51.27 20 513140.36 ABC transporter, ATP-binding Intracellular protein OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0719 PE = 4 SV = 1 -[B2INF2_STRPS] B2IRS2 50.33 12 457 140.22 Cell division protein FtsA OS= Intracellular Streptococcus pneumoniae (strain CGSP14) GN = ftsZ PE =3 SV = 1 - [B2IRS2_STRPS] B2IPY5 51.98 15 379 138.63 Glycogenbiosynthesis protein GlgD Intracellular OS = -Streptococcus pneumoniae(strain CGSP14) GN = glgD PE = 4 SV = 1 - [B2IPY5_STRPS] B2IQH2 47.59 12187 137.62 50S ribosomal protein L10 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = rpIJ PE = 3 SV = 1 - [B2IQH2_STRPS]B2IPM7 29.64 14 577 137.31 Phosphoenolpyruvate-protein Intracellularphosphotransferase OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1121 PE = 3 SV = 1 - [B2IPM7_STRPS] B2ISJ8 57.69 11 156 133.5 30Sribosomal protein S7 OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = rpsG PE = 3 SV = 1 - [RS7_STRPS] B2IS54 74.24 10 132 133.2830S ribosomal protein S8 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = rpsH PE = 3 SV = 1 - [RS8_STRPS] B2INZ1 46.38 17470 132.61 FeS assembly protein SufB OS = Intracellu Streptococcuspneumoniae (strain lar CGSP14) GN = SPCG_0816 PE = 4 SV = 1 -[B2INZ1_STRPS] B2IS53 62.92 10 89 131.91 30S ribosomal protein S14 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpsN PE = 3SV = 1 - [RS14_STRPS] B2IS59 63.7 9 146 131.45 50S ribosomal protein L15OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpIO PE= 3 SV = 1 - [RL15_STRPS] B2INH5 50.5 17 299 127.85 SPFH domain/Band 7family OS = N- Streptococcus pneumoniae (strain terminally CGSP14) GN =SPCG_2124 PE = 4 SV = 1 - anchored [B2INH5_STRPS] (No CS) B2IRE3 73.4212 158 127.54 PTS system, IIB component OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0062 PE = 4 SV = 1 - [B2IRE3_STRPS]B2IN11 64.88 14 299 125.53 UTP--glucose-1-phosphate Secretoryuridylyltransferase OS = (released) Streptococcus pneumoniae (strain(with CS) CGSP14) GN = galU PE = 3 SV = 1 - [B2IN11_STRPS] B2IS19 43.7623 617 125.33 Proline--tRNA ligase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = proS PE = 3 SV = 1 - [SYP_STRPS] B2IPB560.5 12 119 125.17 50S ribosomal protein L20 OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = rpIT PE = 3 SV = 1 -[RL20_STRPS] B2IMN9 40.47 11 425 125.01 Uncharacterized protein OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2034 PE= 4 SV = 1 - [B2IMN9_STRPS] B2IS90 47.86 20 537 123.3 Ribonuclease Y OS= -Streptococcus Intracellular pneumoniae (strain CGSP14) GN = rny PE =3 SV = 1 - [B2IS90_STRPS] B2IPB4 36.36 6 66 122.81 50S ribosomal proteinL35 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =rpml PE = 3 SV = 1 - [RL35_STRPS] B2IQW2 62.61 5 115 121.34 Bacterocintransport accessory Intracellular protein OS = -Streptococcus pneumoniae(strain CGSP14) GN = bta PE = 4 SV = 1 - [B2IQW2_STRPS] B2INS1 30.9 21848 120 Aminopeptidase N OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = pepN PE = 4 SV = 1 - [B2INS1_STRPS] B2IPP1 53.66 9246 116.21 Amino acid ABC transporter, ATP- Intracellular bindingprotein OS = -Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1061 PE= 4 SV = 1 - [B2IPP1_STRPS] B2IMN0 34.94 8 352 116.08 Alcoholdehydrogenase, zinc- Intracellular containing OS = -Streptococcuspneumoniae (strain CGSP14) GN = adh PE = 3 SV = 1 - [B2IMN0_STRPS]B2IRN0 53.52 14 284 114.19 Lipoprotein OS = -Streptococcus Lipidpneumoniae (strain CGSP14) anchored GN = SPCG_0153 PE = 3 SV = 1 -[B2IRN0_STRPS] B2IQZ2 48.66 14 374 110.45 Ribosome-binding ATPase YchFOS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = ychF PE= 3 SV = 1 - [B2IQZ2_STRPS] B2IQ10 41.57 13 344 105.33 Uncharacterizedprotein OS = Multi- Streptococcus pneumoniae (strain transmembraneCGSP14) GN = SPCG_1352 PE = 4 SV = 1 - [B2IQ10_STRPS] B2IS49 60.47 7 86105.08 30S ribosomal protein S17 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = rpsQ PE = 3 SV = 1 - [RS17_STRPS] B2IR0735.97 15 442 104.04 Adenylosuccinate synthetase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = purA PE = 3 SV = 1 -[B2IR07_STRPS] B2IMU7 42.55 6 141 103.8 50S ribosomal protein L11 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpIK PE = 3SV = 1 - [RL11_STRPS] B2ISV9 21.53 9 511 103.52 Choline transporter OS =Multi- Streptococcus pneumoniae (strain transmembrane CGSP14) GN =SPCG_1834 PE = 3 SV = 1 - [B2ISV9_STRPS] B2IQV9 40.56 16 572 103.11Phosphoglucomutase OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = pgm PE = 3 SV = 1 - [B2IQV9_STRPS] B2IR17 52.51 12 339101.9 Ribose-phosphate Intracellular pyrophosphokinase OS =Streptococcus pneumoniae (strain CGSP14) GN = prsA PE = 3 SV = 1 -[B2IR17_STRPS] B2IRQ3 37.38 18 650 101.35 Endopeptidase O OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = pepO PE = 4SV = 1 - [B2IRQ3_STRPS] B2IS63 54.72 6 212 100 Adenylate kinase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = adk PE = 3SV = 1 - [KAD_STRPS] B2IMK3 38.15 13 658 99.61 Transketolase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = tktA PE = 3SV = 1 - [B2IMK3_STRPS] B2IMS2 55.97 12 293 98.56 Fructose-bisphosphatealdolase Intracellular OS = -Streptococcus pneumoniae (strain CGSP14) GN= fba PE = 3 SV = 1 - [B2IMS2_STRPS] B2IMH3 55.78 12 346 96.72Catabolite control protein A OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = ccpA PE = 4 SV = 1 - [B2IMH3_STRPS] B2IPY8 48.31 15474 96.35 Glyceraldehyde-3-phosphate Intracellular dehydrogenase,NADP-dependent OS = -Streptococcus pneumoniae (strain CGSP14) GN = gapNPE = 3 SV = 1 - [B2IPY8_STRPS] B2ILY5 30.81 10 555 96.12 Uncharacterizedprotein OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0438 PE = 4 SV = 1 - [B2ILY5_STRPS] B2IS42 95.92 11 98 95.91 50Sribosomal protein L23 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = rpIW PE = 3 SV = 1 - [RL23_STRPS] B2IM28 43.56 13388 95.69 Protein RecA OS = -Streptococcus Intracellular pneumoniae(strain CGSP14) GN = recA PE = 3 SV = 1 - [RECA_STRPS] B2IRR6 27.44 10266 95.52 Cell division protein DivIVA OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1633 PE = 4 SV = 1 - [B2IRR6_STRPS]B2IS68 64.06 8 128 93.87 50S ribosomal protein L17 OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = rpIQ PE = 3 SV = 1 -[RL17_STRPS] B2IPR2 22.47 13 841 93.61 DNA gyrase subunit A OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = gyrA PE = 3SV = 1 - [B2IPR2_STRPS] B2IM73 29.94 12 521 93.46 Amino acid ABCtransporter, amino Multi- acid-binding protein/permease transmembraneprotein OS = -Streptococcus pneumoniae (strain CGSP14) GN = gInH PE = 3SV = 1 - [B2IM73_STRPS] B2INV5 59.04 7 83 93.23 30S ribosomal proteinS20 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =rpsT PE = 3 SV = 1 - [B2INV5_STRPS] B2IRH9 39.61 21 616 93.09 Capsularpolysaccharide Multi- biosynthesis protein, putative OS = transmembraneStreptococcus pneumoniae (strain CGSP14) GN = SPCG_0098 PE = 4 SV = 1 -[B2IRH9_STRPS] B2INY7 55.13 9 263 92.69 ABC transporter, ATP-bindingIntracellular protein OS = -Streptococcus pneumoniae (strain CGSP14) GN= SPCG_0812 PE = 4 SV = 1 - [B2INY7_STRPS] B2INW3 45.21 14 511 92.64Sugar ABC transporter, ATP-binding Intracellular protein OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0788 PE = 4 SV = 1 -[B2INW3_STRPS] B2IQU4 57.5 9 280 89.47 Oxidoreductase, aldo/ketoIntracellular reductase family OS = -Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1466 PE = 4 SV = 1 - [B2IQU4_STRPS] B2ILR7 36.8 13 48187.34 6-phosphogluconate Intracellular dehydrogenase, decarboxylating OS= -Streptococcus pneumoniae (strain CGSP14) GN = gnd PE = 3 SV = 1 -[B2ILR7_STRPS] B2IRW4 23.85 14 436 87.08 GTPase Der OS = -StreptococcusIntracellular pneumoniae (strain CGSP14) GN = der PE = 3 SV = 1 -[DER_STRPS] B2IQW3 46.76 10 278 86.59 Amino acid ABC transporter, aminoLipid acid-binding protein OS = anchored Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1485 PE = 4 SV = 1 - [B2IQW3_STRPS] B2IPI733.14 8 341 85.78 Iron-compound ABC transporter, Lipid ironcompound-binding protein OS = anchored Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1009 PE = 4 SV = 1 - [B2IPI7_STRPS] B2IS65 58.68 12121 85.63 30S ribosomal protein S13 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = rpsM PE = 3 SV = 1 - [RS13_STRPS] B2ISM459.23 8 130 85.44 30S ribosomal protein S9 OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = rpsI PE = 3 SV = 1 -[RS9_STRPS] B2INT7 37.9 17 752 85.35 ATP-dependent Clp protease, ATP-Intracellular binding subunit ClpE OS = Streptococcus pneumoniae (strainCGSP14) GN = clpE PE = 3 SV = 1 - [B2INT7_STRPS] B2IMU6 41.18 8 23885.34 Uncharacterized protein OS = Lipid Streptococcus pneumoniae(strain anchored CGSP14) GN = SPCG_0590 PE = 4 SV = 1 - [B2IMU6_STRPS]B2IQR1 39.42 15 520 84.59 GMP synthase [glutamine- Intracellularhydrolyzing] OS = -Streptococcus pneumoniae (strain CGSP14) GN = guaA PE= 3 SV = 1 - [GUAA_STRPS] B2IPT6 68.42 8 171 84.01 Galactose-6-phosphateisomerase Intracellular subunit LacB OS = -Streptococcus pneumoniae(strain CGSP14) GN = lacB PE = 3 SV = 1 - [LACB_STRPS] B2INL3 30.25 17810 83.74 ATP-dependent Clp protease, ATP- Intracellular binding subunitOS = -Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2162 PE = 3 SV= 1 - [B2INL3_STRPS] B2IS51 78.22 10 101 83.43 50S ribosomal protein L24OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpIX PE= 3 SV = 1 - [RL24_STRPS] B2INN1 42.77 10 346 83.3 Elongation factor TsOS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = tsf PE= 3 SV = 1 - [EFTS_STRPS] B2IS56 67.18 10 131 82.77 50S ribosomalprotein L18 OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = rpIR PE = 3 SV = 1 - [B2IS56_STRPS] B2ISX8 27.68 8 336 82.74Non-canonical purine NTP Intracellular pyrophosphatase OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1853 PE = 3 SV = 1 -[B2ISX8_STRPS] B2INT0 24.07 12 648 81.85 DNA gyrase subunit B OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = gyrB PE = 3SV = 1 - [B2INT0_STRPS] B2IN71 31.13 15 620 81.8 Elongation factor Tufamily protein Intracellular OS = -Streptococcus pneumoniae (strainCGSP14) GN = SPCG_0638 PE = 4 SV = 1 - [B2IN71_STRPS] B2IPU8 19.19 9 71979.81 Ribonucleoside-diphosphate Intracellular reductase OS =-Streptococcus pneumoniae (strain CGSP14) GN = nrdE PE = 3 SV = 1 -[B2IPU8_STRPS] B2INM4 49.45 9 182 79.8 Ribosomal subunit interfaceprotein Intracellular OS = -Streptococcus pneumoniae (strain CGSP14) GN= SPCG_2173 PE = 4 SV = 1 - [B2INM4_STRPS] B2ISV2 28.57 7 392 79.49Galactokinase OS = -Streptococcus Intracellular pneumoniae (strainCGSP14) GN = gaIK PE = 3 SV = 1 - [B2ISV2_STRPS] B2IMZO 45.21 9 449 77.7Glucose-6-phosphate isomerase Intracellular OS = -Streptococcuspneumoniae (strain CGSP14) GN = pgi PE = 3 SV = 1 - [G6PI_STRPS] B2IRD461.8 6 89 77.46 30S ribosomal protein S15 OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = rpsO PE = 3 SV = 1 -[RS15_STRPS] B2IR65 69.38 11 209 76.49 Uncharacterized protein OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1532 PE= 4 SV = 1 - [B2IR65_STRPS] B2IR90 43.02 4 172 76.3 Non-hemeiron-containing ferritin Intracellular OS = -Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1557 PE = 3 SV = 1 - [B2IR90_STRPS] B2IPY628.16 9 380 75.91 Glucose-1-phosphate Intracellular adenylyltransferaseOS = Streptococcus pneumoniae (strain CGSP14) GN = glgC PE = 3 SV = 1 -[GLGC_STRPS] B2IN68 38.1 7 126 75.25 Uncharacterized protein OS =N-terminally Streptococcus pneumoniae (strain anchored CGSP14) GN =SPCG_0635 PE = 4 SV = 1 - (No CS) [B2IN68_STRPS] B2IR60 97.92 8 96 74.9930S ribosomal protein S6 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = rpsF PE = 3 SV = 1 - [RS6_STRPS] B2IQ02 48.45 7 9774.43 50S ribosomal protein L27 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = rpmA PE = 3 SV = 1 - [RL27_STRPS] B2IM4140.05 10 377 74.15 Uncharacterized protein OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = SPCG_1926 PE = 3 SV = 1 -[B2IM41_STRPS] B2IR71 16.41 12 914 73.99 Cation-transporting ATPase,E1-E2 Multi- family OS = -Streptococcus pneumoniae transmembrane (strainCGSP14) GN = pacL PE = 3 SV = 1 - [B2IR71_STRPS] B2IQV3 35.99 6 28972.94 Glycerol uptake facilitator protein, Multi- putative OS =-Streptococcus transmembrane pneumoniae (strain CGSP14) GN = glpF PE = 3SV = 1 - [B2IQV3_STRPS] B2ILW5 35.95 9 306 72.85 Malonyl CoA-acylcarrier protein Intracellular transacylase OS = -Streptococcuspneumoniae (strain CGSP14) GN = fabD PE = 3 SV = 1 - [B2ILW5_STRPS]B2IRQ6 42.81 10 313 71.49 Manganese ABC transporter, Lipidmanganese-binding adhesion anchored liprotein OS = -Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1623 PE = 3 SV = 1 - [B2IRQ6_STRPS]B2IQX4 60.98 9 164 71.29 ATP synthase subunit b OS = N- Streptococcuspneumoniae (strain terminally CGSP14) GN = atpF PE = 3 SV = 1 - anchored[ATPF_STRPS] (No CS) B2IPT0 26.59 10 628 71.03 Elongation factor 4 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = lepA PE = 3SV = 1 - [B2IPT0_STRPS] B2ISJ1 27.74 11 602 69.46Glutamine--fructose-6-phosphate Intracellular aminotransferase[isomerizing] OS = Streptococcus pneumoniae (strain CGSP14) GN = glmS PE= 3 SV = 1 - [B2ISJ1_STRPS] B2IQU1 23.75 13 678 68.47 Glycine--tRNAligase beta subunit Intracellular OS = -Streptococcus pneumoniae (strainCGSP14) GN = glyS PE = 3 SV = 1 - [SYGB_STRPS] B2INH0 41.42 11 338 68.3Ornithine carbamoyltransferase Intracellular OS = -Streptococcuspneumoniae (strain CGSP14) GN = arcB PE = 3 SV = 1 - [OTC_STRPS] B2IQL461.42 12 267 68.04 Phosphate import ATP-binding Intracellular proteinPstB OS = -Streptococcus pneumoniae (strain CGSP14) GN = pstB PE = 3 SV= 1 - [B2IQL4_STRPS] B2ING9 37.9 10 409 65.94 Arginine deiminase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = arcA PE = 3SV = 1 - [ARCA_STRPS] B2IR87 27.8 9 410 65.69 ATP-dependent Clp proteaseATP- Intracellular binding subunit ClpX OS = Streptococcus pneumoniae(strain CGSP14) GN = clpX PE = 3 SV = 1 - [CLPX_STRPS] B2IPY4 19.29 7477 64.75 Glycogen synthase OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = glgA PE = 3 SV = 1 - [GLGA_STRPS] B2ISY0 36 4 10063.86 UPF0154 protein SPCG_1855 OS = N- Streptococcus pneumoniae (strainterminally CGSP14) GN = SPCG_1855 PE = 3 SV = 1 - anchored[B2ISY0_STRPS] (No CS) B2IQL1 28.04 6 271 63.33 Amino acid ABCtransporter, amino Lipid acid-binding protein OS = anchoredStreptococcus pneumoniae (strain CGSP14) GN = SPCG_1383 PE = 4 SV = 1 -[B2IQL1_STRPS] B2IN60 55 5 60 63.27 50S ribosomal protein L32 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpmF PE = 3SV = 1 - [RL32_STRPS] B2IQK0 22.48 15 872 63.2 Alanine -- tRNA ligase OS= Intracellular Streptococcus pneumoniae (strain CGSP14) GN = alaS PE =3 SV = 1 - [SYA_STRPS] B2INE4 30.43 8 447 62.91 DEAD-box ATP-dependentRNA Intracellular helicase CshB OS = -Streptococcus pneumoniae (strainCGSP14) GN = rheB PE = 3 SV = 1 - [B2INE4_STRPS] B2IS50 59.84 8 12262.57 50S ribosomal protein L14 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = rpIN PE = 3 SV = 1 - [RL14_STRPS] B2IND234.2 8 386 62.51 Branched-chain amino acid ABC Lipid transporter, aminoacid-binding anchored protein OS = -Streptococcus pneumoniae strainCGSP14) GN = SPCG_0699 PE = 4 SV = 1 - [B2IND2_STRPS] B2IRC1 40.12 8 33961.9 UDP-glucose 4-epimerase OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = galE PE = 4 SV = 1 - [B2IRC1_STRPS] B2IR96 77.27 7176 61.33 Adenine phosphoribosyltransferase Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = apt PE = 3 SV = 1 -[B2IR96_STRPS] B2IM46 42.61 9 345 61.11 Uncharacterized protein OS = N-Streptococcus pneumoniae (strain terminally CGSP14) GN = SPCG_1931 PE =4 SV = 1 - anchored [B2IM46_STRPS] (No CS) B21MG8 28.22 8 404 61Aspartate aminotransferase OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = aspC PE = 4 SV = 1 - [B2IMG8_STRPS] B2IQN0 59.06 5127 60.7 Uncharacterized protein OS = N- Streptococcus pneumoniae(strain terminally CGSP14) GN = SPCG_1399 PE = 4 SV = 1 - anchored[B2IQN0_STRPS] (No CS) B2IMQ9 22.52 11 737 60.06 PolyribonucleotideIntracellular nucleotidyltransferase OS = Streptococcus pneumoniae(strain CGSP14) GN = pnp PE = 3 SV = 2 - [PNP_STRPS] B2IQ40 24.39 14 89859.69 Phosphoenolpyruvate carboxylase Intracellular OS = -Streptococcuspneumoniae (strain CGSP14) GN = ppc PE = 3 SV = 1 - [B2IQ40_STRPS]B2IND9 60.43 10 230 59.42 Cell division ABC transporter, ATP-Intracellular binding protein FtsE OS = Streptococcus pneumoniae (strainCGSP14) GN = ftsE PE = 4 SV = 1 - [B2IND9_STRPS] B2IQU5 27.92 10 48059.4 Peptidoglycan N-acetylglucosamine N- deacetylase A OS =-Streptococcus terminally pneumoniae (strain CGSP14) anchored GN = pgdAPE = 4 SV = 1 - (No CS) [B2IQU5_STRPS] B2IQ23 39.06 8 425 59.39Uncharacterized protein OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1195 PE = 4 SV = 1 - [B2IQ23_STRPS] B2INH122.54 6 315 59 Carbamate kinase OS = -Streptococcus Secretory pneumoniae(strain CGSP14) (released) GN = arcC PE = 3 SV = 1 - (with CS)[B2INH1_STRPS] B2IRI7 20.76 11 713 58.73 Uncharacterized protein OS =Multi- Streptococcus pneumoniae (strain transmembrane CGSP14) GN =SPCG_0106 PE = 4 SV = 1 - [B2IRI7_STRPS] B2IMP1 37.86 6 206 56.51Hydrolase, haloacid dehalogenase- Intracellular like family OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2031 PE = 4 SV = 1 -[B2IMP1_STRPS] B2IQH6 45.83 13 312 56.05 Bifunctional methioninesulfoxide N- reductase A/B protein OS = terminally Streptococcuspneumoniae (strain anchored CGSP14) GN = msrA PE = 3 SV = 1 - (No CS)[B2IQH6_STRPS] B2IMB6 28.84 8 378 56.02 Chaperone protein DnaJ OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = dnaJ PE = 3SV = 1 - [B2IMB6_STRPS] B2IS18 32.46 7 419 55.7 Zinc metalloprotease OS= Multi- Streptococcus pneumoniae (strain transmembrane CGSP14) GN = eepPE = 3 SV = 1 - [B2IS18_STRPS] B2IR93 26.64 4 259 55.23 Triosephosphateisomerase OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN= tpiA PE = 3 SV = 1 - [B2IR93_STRPS] B2ILX0 37.58 11 455 54.82Acetyl-CoA carboxylase OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = accC PE = 4 SV = 1 - [B2ILX0_STRPS] B2IS39 65.69 6102 54.76 30S ribosomal protein S10 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = rpsJ PE = 3 SV = 1 - [RS10_STRPS] B2IQZ953.89 9 180 54.16 Hypoxanthine-guanine Intracellularphosphoribosyltransferase OS = Streptococcus pneumoniae (strain CGSP14)GN = hgt PE = 4 SV = 1 - [B2IQZ9_STRPS] B2IMZ6 14.35 10 857 54.16 DNAmismatch repair protein MutS Intracellular OS = -Streptococcuspneumoniae (strain CGSP14) GN = hexA PE = 3 SV = 1 - [B2IMZ6_STRPS]B2IR84 36.49 7 296 53.72 Nucleotide-binding protein IntracellularSPCG_1551 OS = -Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1551PE = 3 SV = 1 - [Y1551_STRPS] B2ISX0 31.78 8 321 53.7 Iron-compound ABCtransporter, Lipid iron-compound-binding protein anchored OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1845 PE = 4 SV = 1 -[B2ISX0_STRPS] B2ILX6 56.45 6 186 52.99 Elongation factor P OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = efp PE = 3SV = 1 - [EFP_STRPS] B2IMZ8 23.45 9 563 52.53 Arginine--tRNA ligase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = argS PE = 3SV = 1 - [SYR_STRPS] B2INY8 20.48 9 420 52.28 Uncharacterized protein OS= Intracellular Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0813PE = 4 SV = 1 - [B2INY8_STRPS] B2IRA3 23.52 7 438 52.2 Pyridinenucleotide-disulfide Intracellular oxidoreductase OS = -Streptococcuspneumoniae (strain CGSP14) GN = merA PE = 3 SV = 1 - [B2IRA3_STRPS]B2IPW2 16.67 6 330 51.71 Acetoin dehydrogenase, E1 Intracellularcomponent, beta subunit, putative OS = -Streptococcus pneumoniae (strainCGSP14) GN = acoB PE = 4 SV = 1 - [B2IPW2_STRPS] B2IRU1 40.52 9 23251.05 Putative N-acetylmannosamine-6- Intracellular phosphate2-epimerase OS = Streptococcus pneumoniae (strain CGSP14) GN = nanE PE =3 SV = 1 - [B2IRU1_STRPS] B2ISY2 5.67 4 705 50.76 Trehalose PTS system,IIABC Multi- components OS = -Streptococcus transmembrane pneumoniae(strain CGSP14) GN = treP PE = 4 SV = 1 - [B2ISY2_STRPS] B2IRX3 29.29 10297 50.56 ABC transporter, ATP-binding Intracellular protein OS =-Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1690 PE = 4 SV = 1 -[B2IRX3_STRPS] B2IQY9 24.72 10 453 50.5 Chromosomal replicationinitiator Intracellular protein DnaA OS = -Streptococcus pneumoniae(strain CGSP14) GN = dnaA PE = 3 SV = 1 - [DNAA_STRPS] B2IMI1 36.61 4183 50.13 Transcription Intracellular termination/antiterminationprotein NusG OS = -Streptococcus pneumoniae (strain CGSP14) GN = nusG PE= 3 SV = 1 - [B2IMI1_STRPS] B2ING0 66.32 5 95 49.96 30S ribosomalprotein S16 OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = rpsP PE = 3 SV = 1 - [RS16_STRPS] B2INE1 11.85 6 726 48.69 PTSsystem, IIABC components OS = Multi- Streptococcus pneumoniae (straintransmembrane CGSP14) GN = ptsG PE = 4 SV = 1 - [B2INE1_STRPS] B2IMN6 339 494 48.52 Threonine synthase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = thrC PE = 4 SV = 1 - [B2IMN6_STRPS]B2ILR1 12.8 8 719 47.81 Penicillin-binding protein 1A OS = N-Streptococcus pneumoniae (strain terminally CGSP14) GN = pbp1A PE = 4 SV= 1 - anchored [B2ILR1_STRPS] (No CS) B2IR57 22.53 8 466 47.14 Coffamily protein/peptidyl-prolyl Intracellular cis-trans isomerase,cyclophilin type OS = -Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1524 PE = 4 SV = 1 - [B2IR57_STRPS] B2IRQ1 10.57 5 653 46.37 PTSsystem IIABC components OS = Multi- Streptococcus pneumoniae (straintransmembrane CGSP14) GN = scrA PE = 4 SV = 1 - [B2IRQ1_STRPS] B2IRE523.91 5 276 46.36 PTS system, IID component OS = Multi- Streptococcuspneumoniae (strain transmembrane CGSP14) GN = SPCG_0064 PE = 4 SV = 1 -[B2IRE5_STRPS] B2IRX0 47.08 8 240 46.21 ABC transporter, ATP-bindingIntracellular protein OS = -Streptococcus pneumoniae (strain CGSP14) GN= SPCG_1687 PE = 4 SV = 1 - [B2IRX0_STRPS] B2IR19 40.73 7 302 45.81Amino acid ABC transporter, Lipid periplasmic amino acid-bindinganchored protein, putative OS = -Streptococcus pneumoniae (strainCGSP14) GN = SPCG_0108 PE = 4 SV = 1 - [B2IR19_STRPS] B2IQL3 29.76 6 25245.8 Phosphate import ATP-binding Intracellular protein PstB OS =-Streptococcus pneumoniae (strain CGSP14) GN = pstB PE = 3 SV = 1 -[B2IQL3_STRPS] B2IQ98 60.64 5 94 45.67 50S ribosomal protein L31 type BIntracellular OS = -Streptococcus pneumoniae (strain CGSP14) GN = rpmEPE = 3 SV = 1 - [B2IQ98_STRPS] B2IRE7 15.8 6 386 44.99 Sugar isomerasedomain protein Intracellular AgaS OS = -Streptococcus pneumoniae (strainCGSP14) GN = agaS PE = 4 SV = 1 - [B2IRE7_STRPS] B2IMZ5 21.99 9 56444.92 ABC transporter, ATP- Multi- binding/permease protein OS =transmembrane Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2042 PE= 4 SV = 1 - [B2IMZ5_STRPS] B2IRQ7 56.4 7 172 44.31 Probable thiolperoxidase OS = Intracellular Streptococcus pneumoniae (strain CGSP14)GN = tpx PE = 3 SV = 1 - [B2IRQ7_STRPS] B2ILX8 33.61 8 488 43.22Glutamyl-tRNA(Gln) Intracellular amidotransferase subunit A OS =Streptococcus pneumoniae (strain CGSP14) GN = gatA PE = 3 SV = 1 -[GATA_STRPS] B2IM45 29.04 8 427 43.07 UDP-N-acetylglucosamine 1-Intracellular carboxyvinyltransferase OS = Streptococcus pneumoniae(strain CGSP14) GN = murA PE = 3 SV = 1 - [B2IM45_STRPS] B2IM60 25 6 41643.06 Diaminopimelate decarboxylase Intracellular OS = -Streptococcuspneumoniae (strain CGSP14) GN = lysA PE = 3 SV = 1 - [B2IM60_STRPS]B2IRA1 21.95 9 524 42.94 DEAD-box ATP-dependent RNA Intracellularhelicase CshA OS = -Streptococcus pneumoniae (strain CGSP14) GN = cshAPE = 3 SV = 1 - [B2IRA1_STRPS] B2IR78 17.33 6 450 42.72Phosphoglucosamine mutase OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = glmM PE = 3 SV = 1 - [GLMM_STRPS] B2IST7 22.06 7417 42.6 Capsular polysaccharide Intracellular biosynthesis protein,putative OS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1812 PE= 3 SV = 1 - [B2IST7_STRPS] B2INT8 57.89 2 76 41.48 Uncharacterizedprotein OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0763 PE = 4 SV = 1 - [B2INT8_STRPS] B2IQ61 25.95 6 289 41.38Choline kinase OS = -Streptococcus Intracellular pneumoniae (strainCGSP14) GN = pck PE = 4 SV = 1 - [B2IQ61_STRPS] B2ILR6 14.66 5 464 41.32Mid-cell-anchored protein Z OS = Multi- Streptococcus pneumoniae (straintransmembrane CGSP14) GN = mapZ PE = 3 SV = 1 - [B2ILR6_STRPS] B2ILR429.91 3 117 40.9 Cell cycle protein GpsB OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = gpsB PE = 3 SV = 1 -[B2ILR4_STRPS] B2IQ25 28.88 8 419 40.5 UDP-N-acetylglucosamine 1-Intracellular carboxyvinyltransferase OS = Streptococcus pneumoniae(strain CGSP14) GN = murZ PE = 3 SV = 1 - [B2IQ25_STRPS] B2IQH8 27.11 8439 40.48 Homoserine dehydrogenase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = hom PE = 3 SV = 1 - [B2IQH8_STRPS]B2IN58 28.96 6 335 40.08 Uncharacterized protein OS = Multi-Streptococcus pneumoniae (strain transmembrane CGSP14) GN = SPCG_2101 PE= 4 SV = 1 - [B2IN58_STRPS] B2IRK5 20.21 7 559 40.01 Ribonuclease J OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = rnj PE = 3SV = 1 - [B2IRK5_STRPS] B2INR2 26.51 10 679 39.94 Methionine--tRNAligase OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =metG PE = 3 SV = 1 - [B2INR2_STRPS] B2IQ29 30.5 8 436 39.93 GTPase ObgOS = -Streptococcus Intracellular pneumoniae (strain CGSP14) GN = obg PE= 3 SV = 1 - [OBG_STRPS] B2IMY2 73.21 7 56 39.82 Uncharacterized proteinOS = Secretory Streptococcus pneumoniae (strain (released) CGSP14) GN =SPCG_0626 PE = 4 SV = 1 - (with CS) [B2IMY2_STRPS] B2IPW3 33.6 8 37539.59 Dihydrolipoamide acetyltransferase Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = acoC PE = 3 SV = 1 -[B2IPW3_STRPS] B2IR18 38.5 6 213 39.57 Amino acid ABC transporter, ATP-Intracellular binding protein, putative OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_0107 PE = 4 SV = 1 - [B2IRI8_STRPS] B2ISR013.08 6 535 39.53 Glucan 1,6-alpha-glucosidase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = dexB PE = 4 SV = 1 -[B2ISR0_STRPS] B2IM72 56.91 6 246 39.3 Amino acid ABC transporter, ATP-Intracellular binding protein OS = -Streptococcus pneumoniae (strainCGSP14) GN = glnQ PE = 4 SV = 1 - [B2IM72_STRPS] B2IS45 48.25 3 11439.27 50S ribosomal protein L22 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = rpIV PE = 3 SV = 1 - [RL22_STRPS] B2IM2626.1 5 318 39.16 Autolysin OS = -Streptococcus Intracellular pneumoniae(strain CGSP14) GN = lytA PE = 4 SV = 1 - [B2IM26_STRPS] B2IM71 46.48 471 38.4 Uncharacterized protein OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0446 PE = 4 SV = 1 - [B2IM71_STRPS]B2INE0 30.09 8 329 38.01 Cell division protein FtsX OS = Multi-Streptococcus pneumoniae (strain transmembrane CGSP14) GN = ftsX PE = 3SV = 1 - [B2INE0_STRPS] B2IQ19 15.99 8 763 37.52 DNA helicase OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = pcrA PE = 3SV = 1 - [B2IQ19_STRPS] B2IRN7 21.52 6 660 37.25 Threonine--tRNA ligaseOS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = thrS PE= 3 SV = 1 - [B2IRN7_STRPS] B2IS67 24.44 7 311 36.89 DNA-directed RNApolymerase Intracellular subunit alpha OS = -Streptococcus pneumoniae(strain CGSP14) GN = rpoA PE = 3 SV = 1 - [B2IS67_STRPS] B2IQA4 21.65 8448 36.16 Glutamate dehydrogenase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = gdhA PE = 3 SV = 1 - [B2IQA4_STRPS]B2INE5 20.96 6 396 36.08 S-adenosylmethionine synthase Intracellular OS= -Streptococcus pneumoniae (strain CGSP14) GN = metK PE = 3 SV = 1 -[METK_STRPS] B2IQN3 51.52 4 66 35.91 30S ribosomal protein S21 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpsU PE = 3SV = 1 - [B2IQN3_STRPS] B2IRK4 18.21 8 637 35.62 tRNA uridine 5-Intracellular carboxymethylaminomethyl modification enzyme MnmG OS =Streptococcus pneumoniae (strain CGSP14) GN = mnmG PE = 3 SV = 1 -[B2IRK4_STRPS] B2IR76 14.89 4 282 35.33 DegV family protein OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1543 PE= 4 SV = 1 - [B2IR76_STRPS] B2IRT0 25.85 4 294 35.15 ROK family proteinOS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1647 PE = 4 SV = 1 - [B2IRT0_STRPS] B2IR21 58.62 7 145 35.06Uncharacterized protein OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = SPCG_0033 PE = 4 SV = 1 - [B2IR21_STRPS] B2IPF425.4 8 311 35.02 Adhesion lipoprotein OS = Lipid Streptococcuspneumoniae (strain anchored CGSP14) GN = Imb PE = 3 SV = 1 -[B2IPF4_STRPS] B2ILV9 26.21 6 454 34.9 Aspartokinase OS = -StreptococcusIntracellular pneumoniae (strain CGSP14) GN = lysC PE = 3 SV = 1 -[B2ILV9_STRPS] B2IPP0 17.37 7 495 34.88 Glucose-6-phosphate 1-Intracellular dehydrogenase OS = -Streptococcus pneumoniae (strainCGSP14) GN = zwf PE = 3 SV = 1 - [B2IPP0_STRPS] B2IRU0 8.43 5 510 34.78PTS system, IIBC components OS = Multi- Streptococcus pneumoniae (straintransmembrane CGSP14) GN = SPCG_1657 PE = 4 SV = 1 - [B2IRU0_STRPS]B2IQX1 34.93 7 292 34.34 ATP synthase gamma chain OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = atpG PE = 3 SV = 1 -[ATPG_STRPS] B2ISZ2 16.94 5 490 34.05 Sucrose phosphorylase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1867 PE= 4 SV = 1 - [B2ISZ2_STRPS] B2IP00 28.21 6 397 33.73 Serine protease OS= -Streptococcus N- pneumoniae (strain CGSP14) terminally GN = sphtra PE= 4 SV = 1 - anchored [B2IP00_STRPS] (No CS) B2INW0 31.82 5 220 33.64Deoxyribose-phosphate aldolase Intracellular OS = -Streptococcuspneumoniae (strain CGSP14) GN = deoC PE = 3 SV = 1 - [DEOC_STRPS] B2IM9815.7 6 535 33.16 CTP synthase OS = -Streptococcus Intracellularpneumoniae (strain CGSP14) GN = pyrG PE = 3 SV = 1 - [B2IM98_STRPS]B2IMU0 11.8 4 466 33.1 Dipeptidase OS = -Streptococcus Intracellularpneumoniae (strain CGSP14) GN = pepV PE = 4 SV = 1 - [B2IMU0_STRPS]B2IPU4 37.15 5 253 33.02 Lactose phosphotransferase system Intracellularrepressor OS = -Streptococcus pneumoniae (strain CGSP14) GN = lacR PE =4 SV = 1 - [B2IPU4_STRPS] B2IRR9 38.55 5 179 32.57 Cell division proteinSepF OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =sepF PE = 3 SV = 1 - [SEPF_STRPS] B2IQJ5 27.56 5 225 32.433-dehydroquinate dehydratase OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = aroD PE = 3 SV = 1 - [AROD_STRPS] B2INC8 38.43 7216 32.37 Uracil phosphoribosyltransferase Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = upp PE = 3 SV = 1 -[B2INC8_STRPS] B2INU6 33.73 8 424 32.09 Phosphopentomutase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = deoB PE = 3SV = 1 - [B2INU6_STRPS] B2IME1 24.87 6 378 32.06 TranscriptionIntracellular termination/antitermination protein NusA OS =-Streptococcus pneumoniae (strain CGSP14) GN = nusA PE = 3 SV = 1 -[B2IME1_STRPS] B2INN5 22.79 4 272 31.93 Cell shape-determining proteinN- MreC OS = -Streptococcus pneumoniae terminally (strain CGSP14) GN =mreC PE = 3 anchored SV = 1 - [B2INN5_STRPS] (No CS) B2IN76 12.22 6 45031.55 UDP-N-acetylmuramoylalanine--D- Intracellular glutamate ligase OS= -Streptococcus pneumoniae (strain CGSP14) GN = murD PE = 3 SV = 1 -[MURD_STRPS] B2INF3 16.1 4 267 31.17 Peptidyl-prolyl cis-trans isomeraseLipid OS = -Streptococcus pneumoniae (strain anchored CGSP14) GN = ppiAPE = 3 SV = 1 - [B2INF3_STRPS] B2IQ22 31.82 6 286 30.97 Methionineaminopeptidase OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = map PE = 1 SV = 1 - [B2IQ22_STRPS] B2IS48 70.59 5 68 30.850S ribosomal protein L29 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = rpmC PE = 3 SV = 1 - [RL29_STRPS] B2IS10 19.57 6649 30.36 DNA mismatch repair protein MutL Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = mutL PE = 3 SV = 1 -[MUTL_STRPS] B2IM44 29.93 6 274 30.16 DNA-entry nuclease OS = N-Streptococcus pneumoniae (strain terminally CGSP14) GN = endA PE = 4 SV= 1 - anchored [B2IM44_STRPS] (No CS) B2INF7 54.46 5 112 29.66 ATP conedomain-containing Intracellular protein OS = -Streptococcus pneumoniae(strain CGSP14) GN = SPCG_0728 PE = 4 SV = 1 - [B2INF7_STRPS] B2IS3413.3 7 737 29.66 Anaerobic ribonucleoside Intracellular triphosphatereductase OS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0212PE = 4 SV = 1 - [B2IS34_STRPS] B2IN20 40.78 3 103 29.58 Uncharacterizedprotein OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =SPCG_2067 PE = 4 SV = 1 - [B2IN20_STRPS] B2IMB7 38.95 4 95 29.51Uncharacterized protein OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = SPCG_0492 PE = 4 SV = 1 - [B2IMB7_STRPS] B2IMY920.58 6 486 29.3 Glutamate--tRNA ligase OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = gltX PE = 3 SV = 1 - [SYE_STRPS] B2IQ7546.77 5 186 29.27 LemA protein OS = -Streptococcus N- pneumoniae (strainCGSP14) terminally GN = lemA PE = 4 SV = 1 - anchored [B2IQ75_STRPS] (NoCS) B21PN9 20.75 6 429 29.19 Signal recognition particle receptorIntracellular FtsY OS = -Streptococcus pneumoniae (strain CGSP14) GN =ftsY PE = 3 SV = 1 - [B2IPN9_STRPS] B2INM2 45.33 7 150 28.94 50Sribosomal protein L9 OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = rplI PE = 3 SV = 1 - [RL9_STRPS] B2IR80 20.89 5 292 28.84Uncharacterized protein OS = Multi- Streptococcus pneumoniae (straintransmembrane CGSP14) GN = SPCG_1547 PE = 4 SV = 1 - [B2IR80_STRPS]B2IPW9 32.16 8 283 28.64 Ribosome biogenesis GTPase A OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = SPCG_1141 PE = 3 SV = 1 -[B2IPW9_STRPS] B2IN61 75.51 4 49 28.27 50S ribosomal protein L33 OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = rpmG PE = 3SV = 1 - [RL33_STRPS] B2IM15 25.95 5 158 28.26 Uncharacterized proteinOS = Multi- Streptococcus pneumoniae (strain transmembrane CGSP14) GN =SPCG_1901 PE = 4 SV = 1 - [B2IM15_STRPS] B2IPZ5 13.9 7 633 28.19 ABCtransporter, ATP-binding Intracellular protein OS = -Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1167 PE = 4 SV = 1 - [B2IPZ5_STRPS]B2IMA9 20.94 6 487 28.12 Type I restriction-modification Intracellularsystem, M subunit OS = Streptococcus pneumoniae (strain CGSP14) GN =hsdM PE = 4 SV = 1 - [B2IMA9_STRPS] B2IQ67 11.63 7 1058 28.04Carbamoyl-phosphate synthase Intracellular large chain OS =-Streptococcus pneumoniae (strain CGSP14) GN = carB PE = 3 SV = 1 -[CARB_STRPS] B2IMV8 16.5 4 491 28.03 PTS system, IIC component, Multi-putative OS = -Streptococcus transmembrane pneumoniae (strain CGSP14) GN= SPCG_0602 PE = 4 SV = 1 - [B2IMV8_STRPS] B2INP6 15.54 4 341 28Tryptophan--tRNA ligase OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = trpS PE = 3 SV = 1 - [B2INP6_STRPS] B2INU9 26.02 4269 27.83 Purine nucleoside phosphorylase Intracellular OS =-Streptococcus pneumoniae (strain CGSP14) GN = pnp PE = 3 SV = 1 -[B2INU9_STRPS] B2IMS9 20.8 7 553 27.26 Ribonuclease J OS =-Streptococcus Intracellular pneumoniae (strain CGSP14) GN = rnj PE = 3SV = 1 - [B2IMS9_STRPS] B2ILY3 41.94 4 62 27.17 50S ribosomal proteinL28 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN =rpmB PE = 3 SV = 1 - [RL28_STRPS] B2IRW8 34.46 5 177 26.92Transcriptional repressor NrdR OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = nrdR PE = 3 SV = 1 - [B2IRW8_STRPS]B2IMH1 20.97 7 472 26.92 Cof family protein OS = -StreptococcusIntracellular pneumoniae (strain CGSP14) GN = SPCG_1962 PE = 4 SV = 1 -[B2IMH1_STRPS] B21PM 6 13.79 2 87 26.87 Phosphocarrier protein HPr OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1120 PE= 4 SV = 1 [B2IPM6_STRPS] B2IRB7 27.22 2 158 26.84 Uncharacterizedprotein OS = N- terminally Streptococcus pneumoniae (strain anchoredCGSP14) GN = SPCG_1584 PE = 4 SV = 1 - (No CS) [B2IRB7_STRPS] B2ILW423.46 5 324 26.69 Enoyl-(Acyl-carrier-protein) Intracellular reductaseOS = -Streptococcus pneumoniae (strain CGSP14) GN = fabK PE = 4 SV = 1 -[B2ILW4_STRPS] B2IN41 16.32 7 429 26.52 Histidine -- tRNA ligase OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = hisS PE = 3SV = 1 - [SYH_STRPS] B2IQU2 16.07 3 305 26.2 Glycine -- tRNA ligasealpha subunit Intracellular OS = -Streptococcus pneumoniae (strainCGSP14) GN = glyQ PE = 3 SV = 1 - [SYGA_STRPS] B2IPW1 23.91 4 322 26.19Acetoin dehydrogenase, E1 Intracellular component, alpha subunit,putative OS = -Streptococcus pneumoniae (strain CGSP14) GN = acoA PE = 4SV = 1 - [B2IPW1_STRPS] B2ISG1 25.79 5 190 26.04 Uncharacterized proteinOS = Multi- Streptococcus pneumoniae (strain transmembrane CGSP14) GN =SPCG_1779 PE = 4 SV = 1 - [B2ISG1_STRPS] B2IPZ7 19.35 4 279 25.42 DegVfamily protein OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1169 PE = 4 SV = 1 - [B2IPZ7_STRPS] B2ISK9 13.48 4 26725.25 PTS system, mannose-specific IIC Multi- component OS =-Streptococcus transmembrane pneumoniae (strain CGSP14) (Lipid GN = manMPE = 4 SV = 1 - modified [B2ISK9_STRPS] N-termini) B2IS70 20.32 5 43325.16 UPF0210 protein SPCG_0246 OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0246 PE = 3 SV = 1 - [B2IS70_STRPS]B2INL6 15.22 4 335 24.97 ABC transporter, substrate-binding Lipidprotein, putative OS = -Streptococcus anchored pneumoniae (strainCGSP14) GN = SPCG_2165 PE = 4 SV = 1 - [B2INL6_STRPS] B2IPF2 46.6 4 19124.92 Thioredoxin family protein OS = Lipid Streptococcus pneumoniae(strain anchored CGSP14) GN = SPCG_0974 PE = 4 SV = 1 - [B2IPF2_STRPS]B2INJ2 24.59 7 427 24.82 DItD protein OS = -Streptococcus N- pneumoniae(strain CGSP14) terminally GN = dltD PE = 4 SV = 1 - anchored[B2INJ2_STRPS] (No CS) B2IPH6 13.65 3 359 24.36 Peptide chain releasefactor 1 OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN= prfA PE = 3 SV = 1 - [RF1_STRPS] B2ILY0 20.43 4 514 24.35 Peptidechain release factor 3 OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = prfC PE = 3 SV = 1 - [RF3_STRPS] B2ILV5 20.88 4 18224.04 Alkyl hydroperoxide reductase Intracellular AhpD OS =-Streptococcus pneumoniae (strain CGSP14) GN = mip PE = 3 SV = 1 -[B2ILV5_STRPS] B2IRX2 21.8 4 399 23.82 Uncharacterized protein OS =Multi- Streptococcus pneumoniae (strain transmembrane CGSP14) GN =SPCG_1689 PE = 4 SV = 1 - [B2IRX2_STRPS] B2IM58 11.36 4 308 23.79Membrane protein insertase YidC Multi- OS = -Streptococcus pneumoniae(strain transmembrane CGSP14) GN = yidC PE = 3 SV = 1 - [B2IM58_STRPS]B2IR27 23.03 4 330 23.79 Phosphate acyltransferase OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = plsX PE = 3 SV = 1 -[PLSX_STRPS] B2IQL7 22.95 4 292 23.74 Phosphate ABC transporter, Lipidphosphate-binding protein, anchored putative OS = -Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1389 PE = 4 SV = 1 - [B2IQL7_STRPS]B2IR86 41.62 7 197 23.54 Probable GTP-binding protein EngB IntracellularOS = -Streptococcus pneumoniae (strain CGSP14) GN = engB PE = 3 SV = 1 -[ENGB_STRPS] B2IP77 21 5 419 23.53 Uncharacterized protein OS =Intracellular Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0896 PE= 4 SV = 1 - [B2IP77_STRPS] B2ISL2 23.6 5 339 23.5 Alcohol dehydrogenaseOS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN = adhP PE= 3 SV = 1 - [B2ISL2_STRPS] B2IPR3 21.05 5 247 23.35 Uncharacterizedprotein OS = N- Streptococcus pneumoniae (strain terminally CGSP14) GN =SPCG_1083 PE = 4 SV = 1 - anchored [B2IPR3_STRPS] (with CS) B2IR97 25 6252 23.12 Methyltransferase, putative OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1564 PE = 4 SV = 1 - [B2IR97_STRPS]B2IS98 24.4 4 209 22.81 Probable nicotinate-nucleotide Intracellularadenylyltransferase OS = Streptococcus pneumoniae (strain CGSP14) GN =nadD PE = 3 SV = 1 - [B2IS98_STRPS] B2IQS3 22.11 4 303 22.58 Thioredoxinreductase OS = Intracellular Streptococcus pneumoniae (strain CGSP14) GN= trxB PE = 3 SV = 1 - [B2IQS3_STRPS] B21PW5 31.61 6 329 22.56Lipoate-protein ligase, putative OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1137 PE = 4 SV = 1 - [B2IPW5_STRPS]B2IMB4 30.77 5 182 22.24 Protein GrpE OS = -Streptococcus Intracellularpneumoniae (strain CGSP14) GN = grpE PE = 3 SV = 1 - [B2IMB4_STRPS]B2ILY4 43.8 3 121 22.23 Uncharacterized protein OS = IntracellularStreptococcus pneumoniae (strain CGSP14) GN = asp23 PE = 4 SV = 1 -[B2ILY4_STRPS] B2IPH3 18.37 7 479 22.18 tRNA modification GTPase MnmEIntracellular OS = -Streptococcus pneumoniae (strain CGSP14) GN = thdFPE = 3 SV = 1 - [B2IPH3_STRPS] B2INP7 17.59 6 540 22.16 ABC transporter,ATP-binding Intracellular protein OS = -Streptococcus pneumoniae (strainCGSP14) GN = SPCG_2196 PE = 4 SV = 1 - [B2INP7_STRPS] B2IP74 14.66 5 49121.9 Lysine decarboxylase OS = Intracellular Streptococcus pneumoniae(strain CGSP14) GN = cad PE = 4 SV = 1 - [B2IP74_STRPS] B2ISG4 52.24 267 21.88 Uncharacterized protein OS = Intracellular Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1782 PE = 3 SV = 1 - [B2ISG4_STRPS]B2ISY1 17.74 7 541 21.79 Dextran glucosidase DexS, putativeIntracellular OS = -Streptococcus pneumoniae (strain CGSP14) GN = dexSPE = 4 SV = 1 - [B2ISY1_STRPS] B2IMA5 15.4 5 448 21.67 Glutaminesynthetase, type I OS = Intracellular Streptococcus pneumoniae (strainCGSP14) GN = gInA PE = 3 SV = 1 - [B2IMA5_STRPS] B2IN77 24.72 6 35221.61 UDP-N-acetylglucosamine -- N- Intracellularacetylmuramyl-(pentapeptide) pyrophosphoryl-undecaprenol N-acetylglucosamine transferase OS = Streptococcus pneumoniae (strainCGSP14) GN = murG PE = 3 SV = 1 - [MURG_STRPS] B2IQ12 25.39 5 319 21.48Ribose-phosphate Intracellular pyrophosphokinase OS = Streptococcuspneumoniae (strain CGSP14) GN = prs PE = 3 SV = 1 - [B2IQ12_STRPS]B2ISL4 16.3 4 270 21.32 Cof family protein OS = -StreptococcusIntracellular pneumoniae (strain CGSP14) GN = SPCG_0299 PE = 4 SV = 1 -[B2ISL4_STRPS] B2IQ78 13.38 5 523 21.2 Signal recognition particleprotein Intracellular OS = -Streptococcus pneumoniae (strain CGSP14) GN= ffh PE = 3 SV = 1 - [B2IQ78_STRPS] B2IS71 21.69 5 249 21.06Phosphoglycerate mutase family Intracellular protein OS = -Streptococcuspneumoniae (strain CGSP14) GN = gpmB PE = 4 SV = 1 - [B2IS71_STRPS]B2IM30 15.38 £ 5 338 20.64 Transcriptional regulator, putativeSecretory) OS = -Streptococcus pneumoniae (released (strain CGSP14) GN =lytR PE = 4 SV = 1 - (with CS) [B2IM30_STRPS] B2IQN9 10.91 4 486 20.01Nicotinate Intracellular phosphoribosyltransferase OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1408 PE = 3 SV = 1 - [B2IQN9_STRPS]

Example 2: Microparticles (MPs) Confer Serotype-Independent ProtectiveImmunity

For each immunization experiment male C57BL/6 wild-type mice ca 5 weeksold were used. Before immunization, mice were anesthetized by inhalationof isofluorane (Abbott) and then intranasally administered with 50μl/mouse of microparticles (MP) combined with the adjuvant aluminiumhydroxide (Sigma Aldrich, 10 mg/ml in PBS) or the adjuvant alone for thecontrol groups. Immunization was repeated again after two weeks from thefirst immunization following the same conditions described above. After4 weeks of immunization, mice were infected by intranasal administrationof 50 μl/mouse of 5×10⁶ CFU, for S. pneumoniae type 1 infectionexperiments (FIG. 7 ), or 10⁶ CFU, for S. pneumoniae type 3 infectionexperiments (FIG. 11 ).

All mice were anesthetized by inhalation of isofluorane prior tochallenge with bacteria. After the infection, clinical symptoms of themice were monitored multiple times per day (in accordance with theethical permit). Blood samples (5 μl/mouse) were taken every day ofinfection and level of bacteremia was assessed by plating serialdilutions of blood samples onto blood-agar plates. Mice that reachedhumane end-points were anesthetized by inhalation of isofluorane andintranasal administered with 100 μl/mouse of the fluorescent markerBacterisense 645 (Perkin Elmer) 30 minutes before the sacrifice. Priorto sacrifice, mice were anesthetized again (isofluorane inhalation).After sacrifice, mice were placed in the IVIS Spectrum Imaging System todetect the fluorescent signal of the bacterial infection. After the IVISimaging, lungs and spleens were collected for further analysis.Bacterial amount in the lungs was assessed by CFU count of bacteriaafter plating serial dilutions of lung homogenates onto blood-agarplates.

Intranasal immunization with microparticles (MP) from a serotype 4 S.pneumoniae increased survival after a challenge with serotype 1 S.pneumoniae (FIG. 7A). The increase in survival was significant also incomparison to immunization with MV_(L). The immunization greatly reducedthe bacterial load in the lungs of the mice surviving the challenge withserotype 1 S. pneumoniae (FIG. 7B). Furthermore, mice immunized withserotype 3 MP and then challenged with serotype 3 bacteria showed 100%protection in the pneumonia model, higher than immunization with thecurrently available vaccine PCV13, which only conferred a protection of75% in the model. Moreover, protection against pneumococcal infectionwas absent in mice lacking B cells, suggesting the involvement of anadaptive immune response with anti-pneumococcal antibodies (FIG. 11 ).

Example 3: Serotype-Independent Binding and Opsonophagocitic Activity ofAntibodies Raised Against Pneumococcal Membrane Particles

Antibodies raised against the inventive MP from serotype 4 as antigenwere reactive against Streptococcus pneumoniae of other serotypes, suchas 1, 6B and 3 (FIG. 9 ). In other words, the MP were able to elicitproduction of serotype-independent antibodies, much more so than MV_(L)(FIG. 9B).

The production of anti-MP antibodies in mice was more efficient (inparticular at time point 2 weeks) when the antigen was administeredtogether with an adjuvant. However, satisfactory results were obtainableeven without an adjuvant (FIG. 8 ). Moreover, antibodies in sera wereable to bind both the encapsulated T4 (TIGR4) and the unencapsulatedisogenic mutant T4R to the same extent, indicating that the antibodyresponse was capsule-independent.

Importantly, the anti-MP antibodies showed opsonophagocitic activity,increasing adhesion of bacteria and killing of internalized bacteria byRAW murine macrophages (FIG. 10 ).

Materials and Methods

Bacterial strains and growth conditions. In this study six strains ofStreptococcus pneumoniae were used: TIGR4 or T4 (serotype 4) (22), itsisogenic mutants lacking the cytotoxin pneumolysin (T4Δply) (23), theautolysin LytA (T4ΔlytA) (3), or the capsule (T4R) (24), ATCC 6301(serotype 1) and I-33 (serotype 3). Bacteria were grown in C+Y medium pH7.9-8.0 (for serotype 4) or THY medium (for serotype 1 and 3) at 37° C.A spectrophotometer (Genesys 20, Thermo Spectronic) was used to followthe growth by monitoring the optical density (OD) at 600 nm.

Isolation and purification of membrane vesicles (MV_(L)) andmicroparticles (MP). For isolation of MV_(L) from a liquid culture,pneumococcal strains (TIGR4, T4Δply or T4ΔlytA) were grown at 37° C. inC+Y medium, pH 7.9-8.0 until OD_(600 nm)=0.9. The culture was thencentrifuged (17,000×g for 30 minutes at 4° C.) to remove bacterial cellsfrom the supernatant. The cell-free supernatant was filtered through a0.22 μm filter (Sarstedt) and centrifuged (120,000×g for 2 hours at 4°C.) to sediment the vesicles. Pellets were washed twice inphosphate-buffered saline (PBS) and resuspended in PBS.

For isolation of microparticles (MP) from bacteria grown on plates, thepneumococcal strains were streaked on blood agar plates and incubatedovernight at 37° C. with 5% CO₂. Bacteria were harvested from plates,resuspended in PBS and MP were pelleted following the same procedure asmentioned above.

Crude MV_(L)/MP preparations were further purified by density gradientcentrifugation using Optiprep™ Density Gradient Medium (Sigma). Pelletedparticle fractions were adjusted to 50% (w/v) Optiprep™ in a totalvolume of 2 ml and overlayed with one fraction of 30% (w/v) Optiprep™ (9ml) followed by a fraction of 5% (w/v) Optiprep™ (3 ml). Gradients werecentrifuged at 250,000×g for 3 hours at 4° C. and the first 4 ml on top,containing the particles, were collected. After 3 washes with PBS(250,000×g for 2 hours at 4° C.), pellets were recovered in PBS andstored at −80° C.

Electron microscopy. To visualize MV_(L) and MP on bacteria, S.pneumoniae T4R was grown in C+Y medium until OD₆₀₀=0.4 at 37° C.Bacteria were harvested by centrifugation for 10 min at 4,000×g, 4° C.and pellets were suspended in 100 μl PBS. Glow discharged carbon coatedgrids (Oxford Instruments, UK) were incubated for 1 min with a drop ofbacterial solution or purified MV_(L)/MP preparation and negativelystained with 2% uranyl acetate in water (7 times for 10 sec). Specimenswere examined on a FEI CM120 microscope operated at 80 kV. Images werecollected with a side mounted camera MegaView III (Olympus Soft Imagingsolutions).

Atomic force microscopy. 5 μl of MP samples isolated from S. pneumoniaewere placed onto freshly cleaved mica (Goodfellow Cambridge Ltd.,Cambridge, United Kingdom). The specimens on the mica were blot driedand placed into a desiccator for at least 2 h. Imaging was performed ona Nanoscope IIIa (Digital Instruments, Santa Barbara) Atomic ForceMicroscope using Tapping Mode with standard silicon cantileversoscillating at resonant frequency (270 to 305 kHz). Images werecollected at a scan rate of 0.8-1.5 Hz, depending on sample number andthe size of the scan. The final images were fitted in both axes andpresented in a surface plot of the height mode.

SDS-PAGE and western blotting. The total amount of proteins in purifiedMV_(L) or MP, lysates of A549 cells or dendritic cells stimulated withMV_(L) or MP were quantified with Pierce™ BCA Protein Assay Kit (LifeTechnologies). Samples containing equal amounts of total protein wereresolved by SDS-PAGE using 4-12% Bis-Tris gels (Life Technologies) andtransferred to PVDF membranes. Membranes were then blocked with 5% skimmilk in PBS containing 0.1% Tween-20 and incubated with antibodies asindicated. For detection of pneumolysin a mouse monoclonal antibody(Abcam, final dilution 1:500) was used. Polyclonal GAPDH (1:2,000) andLytA (1:2,000) (3) antisera raised in rabbits were used. Rabbitpolyclonal antibodies against PspC (1:1,000), RrgB (1:1,000), PsaA(1:25,000), PhtD (1:25,000) and mouse polyclonal antibodies against SrtA(1:500) were kindly provided by Novartis Vaccines and Diagnostics. Asloading control for A549 cells and dendritic cell lysates, a mousemonoclonal β-actin antibody (Santa Cruz) was used as primary antibody.Anti-mouse IgG or anti-rabbit IgG conjugated to horseradish peroxidase(GE Healthcare) were used as secondary antibodies (final dilution1:10,000). Blots were developed with Amersham™ ECL Plus Western blottingdetection system (GE Healthcare Life Sciences), using a ChemiDoc™ XRS+(Bio-Rad Laboratories).

Tandem mass spectrometry. Proteins in MV_(L) or MP samples were reduced,alkylated and in-solution digested by trypsin according to a standardoperating procedure. Thereafter the samples were purified by Pierce C18Spin Columns (Thermo Scientific), dried and resolved in 0.1% formicacid. The resulting peptides were separated in reversed-phase on aC18-column and electrosprayed on-line to a Q Exactive Plus massspectrometer (Thermo Finnigan). Tandem mass spectrometry was performedapplying HCD.

Database searches were made using the Sequest algorithm towards a FASTAdatabase including proteins from Streptococcus pneumoniae TIGR4. Thesearch criteria for protein identification were set to at least twomatching peptides of 95% confidence level per protein. Only proteinswith a Sequest score above 20 were considered for analysis, to avoid thepossibility of false positives.

Subcellular localizations of proteins were predicted using the algorithmof website Locate P located at the following URL:www.cmbi.ru.nl/locatep-db/cgi-bin/locatepdb.py (11).

Hemolysis assay. Purified MV_(L) or MP were incubated in 96-well plateswith blood from buffy coats (diluted 1:50 in PBS containing 1 mMdithiothreitol (DTT)) for 1 hour at 37° C. After 50 minutes 1% TritonX-100 in PBS was added to the positive control wells in order to lyseall the erythrocytes. Plates were then spun at 400×g for 15 minutes at4° C.; supernatant was transferred to an optical plate and the opticaldensity was measured at 540 nm.

A549 cell toxicity assay. A459 lung epithelial cells were grown andmaintained at 37° C., with 5% CO₂ in RPMI medium (Gibco) supplementedwith 10% (v/v) Fetal bovine serum (FBS) (HyClone). To assess MV_(L) orMP associated cytotoxic effects, 0.6×10⁶ A549 cells were seeded in6-well plates and incubated overnight at 37° C. Cells were then washedwith PBS and incubated for 24 hours with medium containing MV_(L) or MPat indicated concentrations. Washed cells were labelled with FixableViability Dye eFluor® 780 (1:50,000, eBioscience) for 30 minutes at 4°C. in the dark, in presence or absence of 0.02% NP40 (Sigma) as positivecontrol, and fixed with 4% paraformaldehyde (PFA) for 30 minutes at roomtemperature.

Next, cells were gently scraped into PBS containing 1% FBS and analyzedin a Gallios™ Flow Cytometer (Beckman Coulter).

A549 cell uptake assay. To assess MV_(L) and MP uptake by A549 cells viaimmunoblotting, 0.6×10⁶ A549 cells were seeded in 6-well plates andincubated overnight at 37° C. Cells were then washed with PBS andincubated for 24 hours with medium containing the particles at indicatedconcentrations. After washing, cells were lysed in RIPA buffercontaining 1× protease inhibitor cocktail (Roche). Cell lysates wereanalyzed by SDS-PAGE and western blotting.

Immunofluorescence microscopy. A549 cells (6.25×10⁴) were seeded in24-well plates with coverslips on the bottom of the wells, and incubatedat 37° C. overnight. Cells were then washed, fixed with 4% PFA for 30minutes and permeabilized with 1% Triton X-100 in PBS for 5 minutes.MV_(L) or MP were detected with mouse monoclonal anti-Ply (1:200) andpolyclonal rabbit antibodies to LytA (1:200). Respective antigens werevisualized with Alexa Fluor® 488-conjugated goat-anti mouse IgG antibody(Life Technologies) and Alexa Fluor® 350-conjugated goat-anti rabbit IgG(Life Technologies) (1:1,000).

Actin cytoskeleton was stained with Alexa Fluor® 594 Phalloidin (LifeTechnologies, 1:40 dilution) for 1 hour and coverslips were mounted withVectashield (Vector Laboratories, Inc.) on microscope slides. Imageswere acquired with a DeltaVision microscope equipped with a60×-objective. Quick projection images of approximately 20 z-stacks weretaken. Orthogonal views were used to visualize the cell monolayer fromthe x, y, and z axes.

After growth, pneumococci (serotypes 1, 3 and 6B) have been stainedusing the sera from the immunized mice as primary antibody (dilution1:100 in PBS 1% BSA) and Alexa Fluor 488 goat anti mouse (dilution 1:500in PBS 1% BSA) as secondary antibody.

Imaging has been performed with high-resolution Delta Vision EliteSystem using 100× objective (1000× total magnification). FITC Laserintensity 50%, exposure time 0,025 ms (FIGS. 13-15 ).

Quantification of the Signal Detected on the Bacteria afterImmunofluorescence Staining

Using the functions Image>Adjust>Threshold and Analyze>Measure ofImageJ, the area covered by the bacteria and the area covered by thesignal detected on the bacteria after staining with sera were selected,defined and measured. The final signal ratio was calculated by dividingthe area of the signal (detected using the sera) by the total area ofthe bacteria.

Isolation and differentiation of human monocyte-derived dendritic cells.Dendritic cells were isolated using RosetteSep™ Human MonocyteEnrichment Cocktail (Stemcell Technologies) according to themanufacturer's instructions. In brief, blood from buffy coats fromhealthy donors was incubated for 20 minutes with RosetteSep™ HumanMonocyte Enrichment Cocktail (Stemcell Technologies), layered on top ofFicoll-Paque™ Plus (GE Healthcare) and centrifuged at 1200×g for 20minutes without acceleration or brake. The monocyte containing layer wasrecovered, cells washed 7 times with PBS and passed through a 100 μmcell strainer. Monocytes were then differentiated for 6 days in RPMIcontaining 10% FBS supplemented with 37.5 ng/ml of Granulocytemacrophage colony-stimulating factor (GM-CSF) (Peprotech) and 37.5 ng/mlof Interleukin 4 (IL-4) (Peprotech), changing the medium after 4 days.For experiments, cells were resuspended in RPMI containing 10% FBS.

Toxicity and apoptosis assay of dendritic cells. 6×10⁵ cells were seededin 96-well plates and incubated with RPMI containing 10% FBS and MV_(L)or MP at indicated concentrations, or with T4R in a multiplicity ofinfection (MOI) of 20, as positive control, for 24 hours. Gentamicin(100 μg/ml, Sigma) was added after 1 hour of incubation to stop bacteriafrom growing in samples stimulated with T4R. Before staining, cells werewashed once in PBS and once in Annexin V Binding Buffer (BD Pharmingen).Staining was performed with Fixable Viability Dye eFluor® 780 (1:50,000,eBioscience) and FITC Annexin V (1:20, BD Pharmingen) for 30 minutes at4° C., followed by two washes with Annexin buffer. Labelled cells werefixed in 4% PFA for 30 minutes, resuspended in PBS containing 1% FBS andanalyzed in a Gallios™ Flow Cytometer (Beckman Coulter).

Activation assay of dendritic cells. 6×10⁵ cells were seeded in 96-wellplates and incubated in RPMI containing 10% FBS with MV_(L) or MP atindicated concentrations, or with 1 μg/ml lipopolysaccharide (LPS)(Sigma) as positive control, for 24 hours. Cells were stained withPhycoerythrin (PE) Mouse Anti-Human CD86 (BD Pharmingen) and PE-Cy™ 5Mouse Anti-Human HLA-DR (BD Pharmingen) for 20 minutes at 4° C., washedtwice with PBS and resuspended in PBS 1% FBS. Labelled cells wereanalyzed in a Gallios™ Flow Cytometer (Beckman Coulter).

Uptake assay with dendritic cells. 10⁶ cells were seeded in 96-wellplates and incubated with RPMI containing 10% FBS, in presence orabsence of the inhibitors cytochalasin D (0.5 μg/ml, Sigma) andwortmannin (0.5 μg/ml, Sigma) (C/W), or methyl-β-cyclodextrin (MβCD) (10μM, Sigma), for 30 minutes. Cells were then incubated with particles atindicated concentration for 1 hour and lysed in RIPA buffer containing1× protease inhibitor cocktail (Roche). Cell lysates were analyzed bySDS-PAGE and western blotting.

Quantification of cytokines. Different cytokines were assessed (IL-6,IL-8, IL-10 and TNF) in cell-free supernatants of 10⁵ dendritic cells byEnzyme-Linked Immunosorbent Assay (ELISA), using commercially availableBD OptEIA™ kits from BD Biosciences. Cells were incubated with particlesat indicated concentrations, or with T4R or LPS (as previouslydescribed), for 24 hours.

Opsonophagocytosis assay with RAW cells. RAW 264.7 murine macrophageswere grown and maintained at 37° C., with 5% CO₂ in RPMI medium (Gibco)supplemented with 10% (v/v) Fetal bovine serum (FBS) (HyClone). Toassess the opsonophagocytosis activity of antibodies in immunized micesera 2×10⁵ RAW 264.7 cells were seeded in 24-well plates and incubatedovernight at 37° C. Serotype 1 bacteria were incubated for 30 minutes at37° C. with 5% CO₂ with 20% serum from mice immunized with MP+adjuvant,or with adjuvant alone as negative control. RAW cells were then washedwith PBS and incubated for 1.5 hours with 2.5×10⁷/well of pre-treatedbacteria. Cells were washed three times with PBS to remove unattachedbacteria. To measure total uptake of bacteria, cells were incubated witha 50/50 solution of 2% saponin (Sigma) and trypsin-EDTA (Gibco) for 15minutes at 37° C., to lyse eukaryotic cells, and total bacteria wereplated for enumeration. To evaluate phagocytosis 300 μg/ml of Gentamicin(Sigma) and 0.12 mg/ml of Penicillin G (Sigma) were added to separatewells and incubated 15 minutes at 37° C., to kill extracellularbacteria. Cells were then washed three times with PBS and incubated witha 50/50 solution of 2% saponin and trypsin-EDTA for 15 minutes at 37° C.to lyse eukaryotic cells. To evaluate killing of bacteria insidemacrophages separate wells were treated with antibiotics (as forphagocytosis), washed three times with PBS and then incubated for 1 hourat 37° C. with medium. Cells were then washed three times with PBS andincubated with a 50/50 solution of 2% saponin and trypsin-EDTA for 15minutes at 37° C. to lyse eukaryotic cells.

Mouse IgG ELISA assay. To detect MV_(L)- and MP-specific mouse IgG insera of immunized mice optical plates (Sarstedt) were coated with 1μg/ml of MP in 0.1 M Sodium Carbonate buffer pH 9.5, overnight at 4° C.Wells were then washed three times with PBS containing 0.05% Tween-20and incubated with PBS with 10% FBS, for 1 hours at room temperature.After three washes wells were incubated with mice sera diluted 1:500 inPBS with 10% FBS, for 2 hours at room temperature. Wells were thenwashed three times and incubated with anti-mouse IgG-HRP (GE Healthcare)diluted 1:500 in PBS 10% FBS, for 2 hours at room temperature. Afterthree washes wells were then incubated with TMB substrate (BDBioscience) for 10 minutes and the reaction was stopped with 1 M H₃PO₄.Absorbance at 450 nm was then measured with a plate reader.

To detect pneumo-specific mouse IgG in sera of immunized mice, bacteriawere grown on blood agar plates overnight at 37° C., resuspended in PBSand heat-inactivated for 2 hours at 60° C. After diluting the bacteriato OD₆₀₀ 0.6, optical plates (Sarstedt) were coated with 100 μl ofbacteria in 0.1 M Sodium Carbonate buffer pH 9.5, overnight at 4° C.Wells were then washed three times with PBS and incubated with PBS with2.5% skim milk, for 2 hours at room temperature. After three washeswells were incubated with mice sera diluted 1:500 in PBS, for 1 hour atroom temperature. Wells were then washed three times and incubated withanti-mouse IgG-HRP (GE Healthcare) diluted 1:500 in PBS, for 1 hour atroom temperature. After three washes well were then incubated with TMBsubstrate (BD Bioscience) for 10 minutes and the reaction was stoppedwith 1 M H₃PO₄. Absorbance at 450 nm was then measured with a platereader.

Statistical analysis. For multiple comparisons the nonparametric ANOVAtest was used to assess the presence of the differences between thegroups, then the Dunn's test was used to make pairwise comparisons; fortwo groups comparison the non-parametric two tailed Wilcoxon's rank sumtest (also known as Mann-Whitney test) was used. Statisticallysignificant data was defined as *P<0.05, **P<0.01, ***P<0.001,****=P<0.0001.

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What is claimed is:
 1. An isolated Streptococcus pneumoniae membranevesicle microparticle (MP), wherein said MP comprises: i. the proteinPly at the level of ≥0.070 μg/μg total protein in the MP; ii. theprotein LytA at the level of ≥0.070 μg/μg total protein in the MP; iii.the protein PspC at the level of ≥0.130 μg/μg total protein in the MP;or iv. the protein RrgB at the level of ≥0.020 μg/μg total protein inthe MP.
 2. The Streptococcus pneumoniae membrane vesicle microparticleaccording to claim 1, further comprising a capsular polysaccharide of acapsular serotype of Streptococcus pneumoniae.
 3. The Streptococcuspneumoniae membrane vesicle microparticle according to claim 1,comprising the protein Ply at the level of ≥0.070 μg/μg total protein inthe MP.
 4. The Streptococcus pneumoniae membrane vesicle microparticleaccording to claim 1, comprising the protein LytA at the level of ≥0.070μg/μg total protein in the MP.
 5. The Streptococcus pneumoniae membranevesicle microparticle according to claim 1, comprising the protein PspCat the level of ≥0.130 μg/μg total protein in the MP.
 6. TheStreptococcus pneumoniae membrane vesicle microparticle according toclaim 1, comprising the protein RrgB at the level of ≥0.020 μg/μg totalprotein in the MP.
 7. The Streptococcus pneumoniae membrane vesiclemicroparticle according to claim 1, comprising the protein PhtD at alevel being at least 2-fold compared to Streptococcus pneumoniaemembrane vesicles obtained in liquid culture in terms of μg/μg totalprotein in the particle or the vesicle, respectively.
 8. TheStreptococcus pneumoniae membrane vesicle microparticle according toclaim 1, comprising the protein RrgA at the level of ≥0.02 μg/μg totalprotein in the MP.
 9. The Streptococcus pneumoniae membrane vesiclemicroparticle according to claim 1, comprising the protein IgA1-proteaseat the level of ≥0.02 μg/μg total protein in the MP.
 10. A compositioncomprising a Streptococcus pneumoniae membrane vesicle microparticlecomprising: i. the protein Ply at the level of ≥0.070 μg/μg totalprotein in the MP; ii. the protein LytA at the level of ≥0.070 μg/μgtotal protein in the MP; iii. the protein PspC at the level of ≥0.130μg/μg total protein in the MP; or iv. the protein RrgB at the level of≥0.020 μg/μg total protein in the MP.
 11. The composition according toclaim 10, wherein the composition is devoid of whole Streptococcuspneumoniae cells.
 12. The composition according to claim 10, furthercomprising capsular polysaccharides from Streptococcus pneumoniae. 13.The composition according to claim 10, comprising Streptococcuspneumoniae membrane vesicle microparticles in an amount of 1 μg/ml. 14.The composition according to claim 13 comprising Streptococcuspneumoniae membrane vesicle microparticles in an amount of 10 μg/ml. 15.The composition according to claim 10, further comprising an adjuvant.16. A method for producing an isolated Streptococcus pneumoniaemicroparticle/s, comprising: a. providing Streptococcus pneumoniae hostcell; b. culturing said host cell under conditions allowing theproduction of Streptococcus pneumoniae membrane vesicle microparticle/saccording to by the host cell; and c. isolating the microparticle/s thusproduced, wherein said microparticles comprise: i. the protein Ply atthe level of ≥0.070 μg/μg total protein in the MP; ii. the protein LytAat the level of ≥0.070 μg/μg total protein in the MP; iii. the proteinPspC at the level of ≥0.130 μg/μg total protein in the MP; or iv. theprotein RrgB at the level of ≥0.020 μg/μg total protein in the MP. 17.The method according to claim 16, wherein the culturing is carried outin solid phase.